Search results for: tumor suppressor gene

Authors: Akulov M. A., Orlova O. R., Zaharov V. O., Tomskij A. A.

Abstract:

Introduction: One of the common postoperative complications of posterior cranial fossa (PCF) and cerebello-pontine angle tumor treatment is a facial nerve palsy, which leads to multiple and resistant to treatment impairments of mimic muscles structure and functions. After 4-6 months after facial nerve palsy with insufficient therapeutic intervention patients develop a postparalythic syndrome, which includes such symptoms as mimic muscle insufficiency, mimic muscle contractures, synkinesis and spontaneous muscular twitching. A novel method of treatment is the use of a recent local neuromuscular blocking agent– botulinum toxin A (BTA). Experience of BTA treatment enables an assumption that it can be successfully used in late facial nerve palsy complications to significantly increase quality of life of patients. Study aim. To evaluate the efficacy of botulinum toxin A (BTA) (Xeomin) treatment in patients with late facial nerve palsy complications. Patients and Methods: 31 patients aged 27-59 years 6 months after facial nerve palsy development were evaluated. All patients received conventional treatment, including massage, movement therapy etc. Facial nerve palsy developed after acoustic nerve tumor resection in 23 (74,2%) patients, petroclival meningioma resection – in 8 (25,8%) patients. The first group included 17 (54,8%) patients, receiving BT-therapy; the second group – 14 (45,2%) patients continuing conventional treatment. BT-injections were performed in synkinesis or contracture points 1-2 U on injured site and 2-4 U on healthy side (for symmetry). Facial nerve function was evaluated on 2 and 4 months of therapy according to House-Brackman scale. Pain syndrome alleviation was assessed on VAS. Results: At baseline all patients in the first and second groups demonstrated аpostparalytic syndrome. We observed a significant improvement in patients receiving BTA after only one month of treatment. Mean VAS score at baseline was 80,4±18,7 and 77,9±18,2 in the first and second group, respectively. In the first group after one month of treatment we observed a significant decrease of pain syndrome – mean VAS score was 44,7±10,2 (р<0,01), whereas in the second group VAS score was as high as 61,8±9,4 points (p>0,05). By the 3d month of treatment pain syndrome intensity continued to decrease in both groups, but, the first group demonstrated significantly better results; mean score was 8,2±3,1 and 31,8±4,6 in the first and second group, respectively (р<0,01). Total House-Brackman score at baseline was 3,67±0,16 in the first group and 3,74±0,19 in the second group. Treatment resulted in a significant symptom improvement in the first group, with no improvement in the second group. After 4 months of treatment House-Brockman score in the first group was 3,1-fold lower, than in the second group (р<0,05). Conclusion: Botulinum toxin injections decrease postparalytic syndrome symptoms in patients with facial nerve palsy.

Keywords: botulinum toxin, facial nerve palsy, postparalytic syndrome, synkinesis

Procedia PDF Downloads 297

Authors: Hossein Rassi, Marjan Moradi fard, Masoud Houshmand

Abstract:

Background: Cervical cancer is multistep disease that is thought to result from an interaction between genetic background and environmental factors. Human Papillomavirus (HPV) infection is the leading risk factor for Cervical Intraepithelial Neoplasia (CIN) and cervical cancer. In other hand, some of p53 and miRNA polymorphism may plays an important role in carcinogenesis. This study attempts to clarify the relation of p53 genotypes and miR-146a rs2910164 polymorphism in cervical lesions. Method: Forty two archival samples with cervical lesion retired from Khatam hospital and 40 sample from healthy persons used as control group. A simple and rapid method was used to detect the simultaneous amplification of the HPV consensus L1 region and HPV-16,-18, -11, -31, 33, and -35 along with the b-globin gene as an internal control. We use Multiplex PCR for detection of P53 and miR-146a rs2910164 genotypes in our lab. Finally, data analysis was performed using the 7 version of the Epi Info(TM) 2012 software and test chi-square(x2) for trend. Results: Cervix lesions were collected from 42 patients with Squamous metaplasia, cervical intraepithelial neoplasia, and cervical carcinoma. Successful DNA extraction was assessed by PCR amplification of b-actin gene (99 bp). According to the results, p53 GG genotype and miR-146a rs2910164 CC genotype was significantly associated with increased risk of cervical lesions in the study population. In this study, we detected 13 HPV 18 from 42 cervical cancer. Conclusion: The connection between several SNP polymorphism and human virus papilloma in rare researches were seen. The reason of these differences in researches' findings can result in different kinds of races and geographic situations and also differences in life grooves in every region. The present study provided preliminary evidence that a p53 GG genotype and miR-146a rs2910164 CC genotype may effect cervical cancer risk in the study population, interacting synergistically with HPV 18 genotype. Our results demonstrate that the testing of p53 codon 72 polymorphism genotypes and miR-146a rs2910164 polymorphism genotypes in combination with HPV18 can serve as major risk factors in the early identification of cervical cancers. Furthermore, the results indicate the possibility of primary prevention of cervical cancer by vaccination against HPV18 in Iran.

Keywords: cervical cancer, miR-146a rs2910164 polymorphism, p53 polymorphism, intraepithelial, neoplasia, HPV

Procedia PDF Downloads 398

Authors: Cecilia Moreira, Rita Paiva, Daniela Macedo, Leonor Ribeiro, Isabel Fernandes, Luis Costa

Abstract:

Background: Head and neck paragangliomas are a rare group of tumors with a large spectrum of clinical manifestations. The approach to evaluate and treat these lesions has evolved over the last years. Surgery was the standard for the approach of these patients, but nowadays new techniques of imaging and radiation therapy changed that paradigm. Despite advances in treating, the growth potential and clinical outcome of individual cases remain largely unpredictable. Objectives: Characterization of our institutional experience with clinical management of these tumors. Methods: This was a cross-sectional study of patients followed in our institution between 01 January and 31 December 2017 with paragangliomas of the head and neck and cranial base. Data on tumor location, catecholamine levels, and specific imaging modalities employed in diagnostic workup, treatment modality, tumor control and recurrence, complications of treatment and hereditary status were collected and summarized. Results: A total of four female patients were followed between 01 January and 31 December 2017 in our institution. The mean age of our cohort was 53 (± 16.1) years. The primary locations were at the level of the tympanic jug (n=2, 50%) and carotid body (n=2, 50%), and only one of the tumors of the carotid body presented pulmonary metastasis at the time of diagnosis. None of the lesions were catecholamine-secreting. Two patients underwent genetic testing, with no mutations identified. The initial clinical presentation was variable highlighting the decrease of visual acuity and headache as symptoms present in all patients. In one of the cases, loss of all teeth of the lower jaw was the presenting symptomatology. Observation with serial imaging, surgical extirpation, radiation, and stereotactic radiosurgery were employed as treatment approaches according to anatomical location and resectability of lesions. As post-therapeutic sequels the persistence of tinnitus and disabling pain stands out, presenting one of the patients neuralgia of the glossopharyngeal. Currently, all patients are under regular surveillance with a median follow up of 10 months. Conclusion: Ultimately, clinical management of these tumors remains challenging owing to heterogeneity in clinical presentation, the existence of multiple treatment alternatives, and potential to cause serious detriment to critical functions and consequently interference with the quality of life of the patients.

Keywords: clinical outcomes, head and neck, management, paragangliomas

Procedia PDF Downloads 144

Authors: German Hernandez-Alonso, Antonio Lazcano, Arturo Becerra

Abstract:

Until today, viruses remain controversial and poorly understood; about their origin, this problem represents an enigma and one of the great challenges for the contemporary biology. Three main theories have tried to explain the origin of viruses: regressive evolution, escaped host gene, and pre-cellular origin. Under the perspective of the escaped host gene theory, it can be assumed a cellular origin of viral components, like protein RNA-binding domains. These universal distributed RNA-binding domains are related to the RNA metabolism processes, including transcription, processing, and modification of transcripts, translation, RNA degradation and its regulation. In the case of viruses, these domains are present in important viral proteins like helicases, nucleases, polymerases, capsid proteins or regulation factors. Therefore, they are implicated in the replicative cycle and parasitic processes of viruses. That is why it is possible to think that those domains present low levels of divergence due to selective pressures. For these reasons, the main goal for this project is to create a catalogue of the RNA-binding domains found in all the available viral proteomes, using bioinformatics tools in order to analyze its evolutionary process, and thus shed light on the general virus evolution. ProDom database was used to obtain larger than six thousand RNA-binding domain families that belong to the three cellular domains of life and some viral groups. From the sequences of these families, protein profiles were created using HMMER 3.1 tools in order to find distant homologous within greater than four thousand viral proteomes available in GenBank. Once accomplished the analysis, almost three thousand hits were obtained in the viral proteomes. The homologous sequences were found in proteomes of the principal Baltimore viral groups, showing interesting distribution patterns that can contribute to understand the evolution of viruses and their host-virus interactions. Presence of cellular RNA-binding domains within virus proteomes seem to be explained by closed interactions between viruses and their hosts. Recruitment of these domains is advantageous for the viral fitness, allowing viruses to be adapted to the host cellular environment.

Keywords: bioinformatics tools, distant homology, RNA-binding domains, viral evolution

Procedia PDF Downloads 387

Authors: Wiam El Kheir, Anais Dumais, Maude Beaudoin, Bernard Marcos, Nick Virgilio, Benoit Paquette, Nathalie Faucheux, Marc-Antoine Lauzon

Abstract:

The high infiltrative pattern of glioblastoma multiforme cells (GBM) is the main cause responsible for the actual standard treatments failure. The tumor high heterogeneity, the interstitial fluid flow (IFF) and chemokines guides GBM cells migration in the brain parenchyma resulting in tumor recurrence. Drug delivery systems emerged as an alternative approach to develop effective treatments for the disease. Some recent studies have proposed to harness the effect CXC-lchemokine-ligand-12 to direct and control the cancer cell migration through delivery system. However, the dynamics of the brain environment on the delivery system remains poorly understood. Nanoparticles (NPs) and hydrogels are known as good carriers for the encapsulation of different agents and control their release. We studied the release of CXCL12 (free or loaded into NPs) from an alginate-based hydrogel under static and indirect perfusion (IP) conditions. Under static conditions, the main phenomena driving CXCL12 release from the hydrogel was diffusion with the presence of strong interactions between the positively charged CXCL12 and the negatively charge alginate. CXCL12 release profiles were independent from the initial mass loadings. Afterwards, we demonstrated that the release could tuned by loading CXCL12 into Alginate/Chitosan-Nanoparticles (Alg/Chit-NPs) and embedded them into alginate-hydrogel. The initial burst release was substantially attenuated and the overall cumulative release percentages of 21%, 16% and 7% were observed for initial mass loadings of 0.07, 0.13 and 0.26 µg, respectively, suggesting stronger electrostatic interactions. Results were mathematically modeled based on Fick’s second law of diffusion framework developed previously to estimate the effective diffusion coefficient (Deff) and the mass transfer coefficient. Embedding the CXCL12 into NPs decreased the Deff an order of magnitude, which was coherent with experimental data. Thereafter, we developed an in-vitro 3D model that takes into consideration the convective contribution of the brain IFF to study CXCL12 release in an in-vitro microenvironment that mimics as faithfully as possible the human brain. From is unique design, the model also allowed us to understand the effect of IP on CXCL12 release in respect to time and space. Four flow rates (0.5, 3, 6.5 and 10 µL/min) which may increase CXCL12 release in-vivo depending on the tumor location were assessed. Under IP, cumulative percentages varying between 4.5-7.3%, 23-58.5%, 77.8-92.5% and 89.2-95.9% were released for the three initial mass loadings of 0.08, 0.16 and 0.33 µg, respectively. As the flow rate increase, IP culture conditions resulted in a higher release of CXCL12 compared to static conditions as the convection contribution became the main driving mass transport phenomena. Further, depending on the flow rate, IP had a direct impact on CXCL12 distribution within the simulated brain tissue, which illustrates the importance of developing such 3D in-vitro models to assess the efficiency of a delivery system targeting the brain. In future work, using this very model, we aim to understand the impact of the different phenomenon occurring on GBM cell behaviors in response to the resulting chemokine gradient subjected to various flow while allowing them to express their invasive characteristics in an in-vitro microenvironment that mimics the in-vivo brain parenchyma.

Keywords: 3D culture system, chemokines gradient, glioblastoma multiforme, kinetic release, mathematical modeling

Procedia PDF Downloads 85

Authors: H. Haseena Banu, D. Karthick, R. Stalin, E. Nandha Kumar, T. P. Sachidanandam, P. Shanthi

Abstract:

Background of the study: Type 2 diabetes is emerging as the predominant metabolic disorder in the world among adults characterized mainly by the resistance of the insulin sensitive tissues towards insulin followed by the decrease in the insulin secretion. The treatment for this disease usually involves treatment with oral synthetic drugs which are known to cause several side effects. Therefore, identification of new biomarkers as therapeutic target is the need of the hour. miRNAs are small, non–protein-coding RNAs that negatively regulate gene expression by promoting degradation and/or inhibit the translation of target mRNAs and have emerged as biomarkers in predicting diabetes mellitus. Objective of the study: To elucidate the therapeutic role of gallic acid in modulating the alterations in glucose metabolism induced by miRNAs 194 and 135a in Type 2 diabetic rats. Materials and Methods: T2D was induced in rats by feeding them with a high fat diet for 2 weeks followed by intraperitoneal injection of 35 mg/kg/body weight (b.wt.) of streptozotocin. Microarrays were used to assess the expression of miRNAs in control, diabetic and gallic acid treated rats. Gene expression studies were carried out by RT PCR analysis. Results: Forty one miRNAs were differentially expressed in Type 2 diabetic rats. Among these, the expression of miRNA 194 was significantly decreased whereas miRNA 135a was significantly increased in Type 2 diabetic rats. The glucose metabolism was also altered significantly in skeletal muscle of Type 2 diabetic rats. Conclusion: T2D is associated with alterations in the expression of miRNAs in skeletal muscle. Both these miRNAs 194 and 135a play an important role in glucose metabolism in skeletal muscle of diabetic rats. Gallic acid effectively ameliorated the alterations in glucose metabolism. Hence, both these miRNAs can serve as biomarkers and therapeutic targets in diabetes mellitus. The study also establishes the role of gallic acid as therapeutic agent. Acknowledgment: The financial assistance provided in the form of ICMR women scientist by ICMR DHR INDIA is gratefully acknowledged here.

Keywords: gallic acid, high fat diet, type 2 diabetes mellitus, miRNAs

Procedia PDF Downloads 349

Authors: Salman Akrokayan

Abstract:

Synthetic cDNA (ScDNA) of granulocyte colony-stimulating factor (G-CSF) was constructed using a DNA synthesizer with the aim to increase its expression level. 5' end of the ScDNA of G-CSF coding region was modified by decreasing the GC content without altering the predicted amino acids sequence. The identity of the resulting protein from ScDNA was confirmed by the highly specific enzyme-linked immunosorbent assay. In conclusion, a synthetic G-CSF cDNA in combination with the recombinant DNA protocol offers a rapid and reliable strategy for synthesizing the target protein. However, the commercial utilization of this methodology requires rigorous validation and quality control.

Keywords: synthetic cDNA, recombinant G-CSF, cloning, gene expression

Procedia PDF Downloads 284

Authors: Yuanyuan Zhang, Yujuan Zheng, Giuseppina Barutello, Sumako Kameishi, Kungchun Chiu, Katharina Hennig, Martial Balland, Federica Cavallo, Lars Holmgren

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Angiogenesis describes that new blood vessels migrate from pre-existing ones to form 3D lumenized structure and remodeling. During directional migration toward the gradient of pro-angiogenic factors, the endothelial cells, especially the tip cells need filopodia to sense the environment and exert the pulling force. Of particular interest are the integrin proteins, which play an essential role in focal adhesion in the connection between migrating cells and extracellular matrix (ECM). Understanding how these biomechanical complexes orchestrate intrinsic and extrinsic forces is important for our understanding of the underlying mechanisms driving angiogenesis. We have previously identified Angiomotin (Amot), a member of Amot scaffold protein family, as a promoter for endothelial cell migration in vitro and zebrafish models. Hence, we established inducible endothelial-specific Amot knock-out mice to study normal retinal angiogenesis as well as tumor angiogenesis. We found that the migration ratio of the blood vessel network to the edge was significantly decreased in Amotec- retinas at postnatal day 6 (P6). While almost all the Amot defect tip cells lost migration advantages at P7. In consistence with the dramatic morphology defect of tip cells, there was a non-autonomous defect in astrocytes, as well as the disorganized fibronectin expression pattern correspondingly in migration front. Furthermore, the growth of transplanted LLC tumor was inhibited in Amot knockout mice due to fewer vasculature involved. By using MMTV-PyMT transgenic mouse model, there was a significantly longer period before tumors arised when Amot was specifically knocked out in blood vessels. In vitro evidence showed that Amot binded to beta-actin, Integrin beta 1 (ITGB1), Fibronectin, FAK, Vinculin, major focal adhesion molecules, and ITGB1 and stress fibers were distinctly induced by Amot transfection. Via traction force microscopy, the total energy (force indicater) was found significantly decreased in Amot knockdown cells. Taken together, we propose that Amot is a novel partner of the ITGB1/Fibronectin protein complex at focal adhesion and required for exerting force transition between endothelial cell and extracellular matrix.

Keywords: angiogenesis, angiomotin, endothelial cell migration, focal adhesion, integrin beta 1

Procedia PDF Downloads 238

Authors: N. D'Amico, E. Grossi, B. Colombo, F. Rigiroli, M. Buscema, D. Fazzini, G. Cornalba, S. Papa

Abstract:

Purpose: To characterize, through a radiomic approach, the nature of nodules considered non-specific by expert radiologists, recognized in magnetic resonance mammography (MRm) with T1-weighted (T1w) sequences with paramagnetic contrast. Material and Methods: 47 cases out of 1200 undergoing MRm, in which the MRm assessment gave uncertain classification (non-specific nodules), were admitted to the study. The clinical outcome of the non-specific nodules was later found through follow-up or further exams (biopsy), finding 35 benign and 12 malignant. All MR Images were acquired at 1.5T, a first basal T1w sequence and then four T1w acquisitions after the paramagnetic contrast injection. After a manual segmentation of the lesions, done by a radiologist, and the extraction of 150 radiomic features (30 features per 5 subsequent times) a machine learning (ML) approach was used. An evolutionary algorithm (TWIST system based on KNN algorithm) was used to subdivide the dataset into training and validation test and to select features yielding the maximal amount of information. After this pre-processing, different machine learning systems were applied to develop a predictive model based on a training-testing crossover procedure. 10 cases with a benign nodule (follow-up older than 5 years) and 18 with an evident malignant tumor (clear malignant histological exam) were added to the dataset in order to allow the ML system to better learn from data. Results: NaiveBayes algorithm working on 79 features selected by a TWIST system, resulted to be the best performing ML system with a sensitivity of 96% and a specificity of 78% and a global accuracy of 87% (average values of two training-testing procedures ab-ba). The results showed that in the subset of 47 non-specific nodules, the algorithm predicted the outcome of 45 nodules which an expert radiologist could not identify. Conclusion: In this pilot study we identified a radiomic approach allowing ML systems to perform well in the diagnosis of a non-specific nodule at MR mammography. This algorithm could be a great support for the early diagnosis of malignant breast tumor, in the event the radiologist is not able to identify the kind of lesion and reduces the necessity for long follow-up. Clinical Relevance: This machine learning algorithm could be essential to support the radiologist in early diagnosis of non-specific nodules, in order to avoid strenuous follow-up and painful biopsy for the patient.

Keywords: breast, machine learning, MRI, radiomics

Procedia PDF Downloads 267

Authors: Anil Kumar, Diwakar Aggarwal, M. Sudhakara Reddy

Abstract:

The wood of Eucalyptus, Populus and bamboos are some important species used as raw material for the manufacture of pulp. However, higher levels of lignin pose a problem during Kraft pulping and yield of pulp is also lower. In order to increase the yield of pulp per unit wood and reduce the use of chemicals during kraft pulping it is important to reduce the lignin content and/or increase cellulose content in wood. Cellulose biosynthesis in wood takes place by the coordinated action of many enzymes. The two important enzymes are KORRIGAN and SUCROSE SYNTHASE. KORRIGAN (Endo-1,4--glucanase) is implicated in the process of editing growing cellulose chains and improvement of the crystallinity of produced cellulose, whereas SUCROSE SYNTHASE is involved in providing substrate (UDP-glucose) for growing cellulose chains. The present study was aimed at the cloning, characterization and overexpression of these genes in Eucalyptus and Populus. An efficient shoot organogenesis protocol from leaf explants taken from micro shoots of the species has been developed. Agrobacterium mediated genetic transformation using Agrobacterium tumefaciens strain EHA105 and LBA4404 harboring binary vector pBI121 was achieved. Both the genes were cloned from cDNA library of Populus deltoides. These were subsequently characterized using various bioinformatics tools. The cloned genes were then inserted into pBI121 under the CaMV35S promotors replacing GUS gene. The constructs were then mobilized into above strains of Agrobacterium and used for the transformation work. Subsequently, genetic transformation of these clones with target genes following already developed protocol is in progress. Four transgenic lines of Eucalyptus tereticornis overexpressing Korrigan gene under the strong constitutive promoters CaMV35S have been developed, which are being further evaluated. Work on development of more transgenic lines overexpressing these genes in Populus and Eucalyptus is also in progress. This presentation will focus on important developments in this direction.

Keywords: Eucalyptus tereticornis, genetic transformation, Kraft pulping Populus deltoides

Procedia PDF Downloads 140

Authors: Sona Babu Rathinam, Lavanya Dharmendran, Therraddi Mutthu

Abstract:

Objectives: The purpose of this paper is to provides an overview of the neuroendocrine tumors that arise in the oral cavity. Material and Methods: An overview of the relevant papers on neuroendocrine tumors of the oral cavity by various authors was studied and summarized. Results: On the basis of the relevant studies, this paper provides an overview of the classification and histological differentiation of the neuroendocrine tumors that arise in the oral cavity. Conclusions: The basis of classification of neuroendocrine tumors is largely determined by their histologic differentiation. Though they reveal biologic heterogeneity, there should be an awareness of the occurrence of such lesions in the oral cavity to enable them to be detected and treated early.

Keywords: malignant peripheral nerve sheath tumor, olfactory neuroblastoma, paraganglioma, schwannoma

Procedia PDF Downloads 80

Authors: Hossein Rassi, Marjan Moradi Fard, Masoud Houshmand

Abstract:

Background: Cervical cancer is multistep disease that is thought to result from an interaction between genetic background and environmental factors. Human papillomavirus (HPV) infection is the leading risk factor for cervical intraepithelial neoplasia (CIN) and cervical cancer. In other hand, some of p53 and miRNA polymorphism may plays an important role in carcinogenesis. This study attempts to clarify the relation of p53 genotypes and miR-146a rs2910164 polymorphism in cervical lesions. Method: Forty two archival samples with cervical lesion retired from Khatam hospital and 40 sample from healthy persons used as control group. A simple and rapid method was used to detect the simultaneous amplification of the HPV consensus L1 region and HPV-16,-18, -11, -31, 33 and -35 along with the b-globin gene as an internal control. We use Multiplex PCR for detection of P53 and miR-146a rs2910164 genotypes in our lab. Finally, data analysis was performed using the 7 version of the Epi Info(TM) 2012 software and test chi-square(x2) for trend. Results: Cervix lesions were collected from 42 patients with Squamous metaplasia, cervical intraepithelial neoplasia, and cervical carcinoma. Successful DNA extraction was assessed by PCR amplification of b-actin gene (99bp). According to the results, p53 GG genotype and miR-146a rs2910164 CC genotype was significantly associated with increased risk of cervical lesions in the study population. In this study, we detected 13 HPV 18 from 42 cervical cancer. Conclusion: The connection between several SNP polymorphism and human virus papilloma in rare researches were seen. The reason of these differences in researches' findings can result in different kinds of races and geographic situations and also differences in life grooves in every region. The present study provided preliminary evidence that a p53 GG genotype and miR-146a rs2910164 CC genotype may effect cervical cancer risk in the study population, interacting synergistically with HPV 18 genotype. Our results demonstrate that the testing of p53 codon 72 polymorphism genotypes and miR-146a rs2910164 polymorphism genotypes in combination with HPV18 can serve as major risk factors in the early identification of cervical cancers. Furthermore, the results indicate the possibility of primary prevention of cervical cancer by vaccination against HPV18 in Iran.

Keywords: cervical cancer, HPV18, p53 codon 72 polymorphism, miR-146a rs2910164 polymorphism

Procedia PDF Downloads 457

Authors: D. Pradhan, G. Tripathy, S. Pradhan

Abstract:

Objectives: Imperviousness gainst estrogen treatments is a noteworthy reason for infection backslide and mortality in estrogen receptor alpha (ERα)- positive breast diseases. Tamoxifen or estrogen withdrawal builds the reliance of breast malignancy cells on INT3 flagging. Here, we researched the commitment of Quercetin and INT3 motioning in endocrine-safe breast tumor cells. Methods: We utilized two models of endocrine treatments safe (ETR) breast tumor: Tamoxifen-safe (TamR) and long haul estrogen-denied (LTED) MCF7 cells. We assessed the transitory and intrusive limit of these cells by Transwell cells. Articulation of epithelial to mesenchymal move (EMT) controllers and in addition INT3 receptors and targets were assessed by constant PCR and western smudge investigation. Besides, we tried in-vitro hostile to Quercetin monoclonal Antibodies (mAbs) and Gamma Secretase Inhibitors (GSIs) as potential EMT inversion remedial specialists. At last, we created stable Quercetin overexpressing MCF7 cells and assessed their EMT components and reaction to Tamoxifen. Results: We found that ETR cells procured an Epithelial to Mesenchymal move (EMT) phenotype and showed expanded levels of Quercetin and INT3 targets. Interestingly, we distinguished more elevated amount of INT3 however lower levels of INT1 and INT3 proposing a change to motioning through distinctive INT3 receptors after obtaining of resistance. Against Quercetin monoclonal antibodies and the GSI PF03084014 were powerful in obstructing the Quercetin/INT3 pivot and in part repressing the EMT process. As a consequence of this, cell relocation and attack were weakened and the immature microorganism like populace was essentially decreased. Hereditary hushing of Quercetin and INT3 prompted proportionate impacts. At long last, stable overexpression of Quercetin was adequate to make MCF7 lethargic to Tamoxifen by INT3 initiation. Conclusions: ETR cells express abnormal amounts of Quercetin and INT3, whose actuation eventually drives intrusive conduct. Hostile to Quercetin mAbs and GSI PF03084014 lessen articulation of EMT particles decreasing cell obtrusiveness. Quercetin overexpression instigates Tamoxifen resistance connected to obtaining of EMT phenotype. Our discovering propose that focusing on Quercetin and INT3 warrants further clinical Correlation as substantial restorative methodologies in endocrine-safe breast.

Keywords: endocrine, epithelial, mesenchymal, INT3, quercetin, MCF7

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Authors: Hana Barak, Alex Sivan, Ariel Kushmaro

Abstract:

Microorganisms are the most diverse and abundant life forms on Earth and account for a large portion of the Earth’s biomass and biodiversity. To date though, our knowledge regarding microbial life is lacking, as it is based mainly on information from cultivated organisms. Indeed, microbiologists have borrowed from astrophysics and termed the ‘uncultured microbial majority’ as ‘microbial dark matter’. The realization of how diverse and unexplored microorganisms are, actually stems from recent advances in molecular biology, and in particular from novel methods for sequencing microbial small subunit ribosomal RNA genes directly from environmental samples termed next-generation sequencing (NGS). This has led us to use NGS that generates several gigabases of sequencing data in a single experimental run, to identify and classify environmental samples of microorganisms. In metagenomics sequencing analysis (both 16S and shotgun), sequences are compared to reference databases that contain only small part of the existing microorganisms and therefore their taxonomy assignment may reveal groups of unknown microorganisms or origins. These unknowns, or the ‘microbial sequences dark matter’, are usually ignored in spite of their great importance. The goal of this work was to develop an improved bioinformatics method that enables more complete analyses of the microbial communities in numerous environments. Therefore, NGS was used to identify previously unknown microorganisms from three different environments (industrials wastewater, Negev Desert’s rocks and water wells at the Arava valley). 16S rRNA gene metagenome analysis of the microorganisms from those three environments produce about ~4 million reads for 75 samples. Between 0.1-12% of the sequences in each sample were tagged as ‘Unassigned’. Employing relatively simple methodology for resequencing of original gDNA samples through Sanger or MiSeq Illumina with specific primers, this study demonstrates that the mysterious ‘Unassigned’ group apparently contains sequences of candidate phyla. Those unknown sequences can be located on a phylogenetic tree and thus provide a better understanding of the ‘sequences dark matter’ and its role in the research of microbial communities and diversity. Studying this ‘dark matter’ will extend the existing databases and could reveal the hidden potential of the ‘microbial dark matter’.

Keywords: bacteria, bioinformatics, dark matter, Next Generation Sequencing, unknown

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Authors: Dong-An Wang

Abstract:

All kinds of microspheres have been extensively employed as carriers for drug, gene and therapeutic cell delivery. Most therapeutic cell delivery microspheres rely on a two-step methodology: fabrication of microspheres and subsequent seeding of cells onto them. In this study, we have developed a novel one-step cell encapsulation technique using a convenient and instant water-in-oil single emulsion approach to form cell-encapsulated gelatin microspheres. This technology is adopted for hyaline cartilage tissue engineering, in which autologous chondrocytes are used as therapeutic cells. Cell viability was maintained throughout and after the microsphere formation (75-100 µm diameters) process that avoids involvement of any covalent bonding reactions or exposure to any further chemicals. Further encapsulation of cell-laden microspheres in alginate gels were performed under 4°C via a prompt process. Upon the formation of alginate constructs, they were immediately relocated into CO2 incubator where the temperature was maintained at 37°C; under this temperature, the cell-laden gelatin microspheres dissolved within hours to yield similarly sized cavities and the chondrocytes were therefore suspended within the cavities inside the alginate gel bulk. Hence, the gelatin cell-laden microspheres served two roles: as cell delivery vehicles which can be removable through temperature curing, and as porogens within an alginate hydrogel construct to provide living space for cell growth and tissue development as well as better permeability for mutual diffusions. These cell-laden microspheres, namely “temperature-cured dissolvable gelatin microsphere based cell carriers” (tDGMCs), were further encapsulated in a chondrocyte-laden alginate scaffold system and analyzed by WST-1, gene expression analyses, biochemical assays, histology and immunochemistry stains. The positive results consistently demonstrated the promise of tDGMC technology in delivering these non-anchorage dependent cells (chondrocytes). It can be further conveniently translated into delivery of other non-anchorage dependent cell species, including stem cells, progenitors or iPS cells, for regeneration of tissues in internal organs, such as engineered hepatogenesis or pancreatic regeneration.

Keywords: biomaterials, tissue engineering, microsphere, hydrogel, porogen, anchorage dependence

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Authors: Sandra Pietri, Helene Dubout, Sabrina Ena, Candice Hoste, Enrico Bastianelli

Abstract:

Bone Therapeutics is a bone cell therapy company addressing high unmet medical needs in the field of bone fracture repair, more specifically in non-union and delayed-union fractures where the bone repair process is impaired. The company has developed a unique allogeneic osteoblastic cell product (ALLOB®) derived from bone marrow which is currently tested in humans in the indication of delayed-union fractures. The purpose of our study was to directly compare ALLOB® vs. non-differentiated mesenchymal stem cells (MSC) for their in vitro osteogenic characteristics and their in vivo osteogenic potential in order to determine which cellular type would be the most adapted for bone fracture repair. Methods: Healthy volunteers’ bone marrow aspirates (n=6) were expended (i) into BM-MSCs using a complete MSC culture medium or (ii) into ALLOB® cells according to its manufacturing process. Cells were characterized in vitro by morphology, immunophenotype, gene expression and differentiation potential. Additionally, their osteogenic potential was assessed in vivo in the subperiosteal calvaria bone formation model in nude mice. Results: The in vitro side-by-side comparison studies showed that although ALLOB® and BM-MSC shared some common general characteristics such as the 3 minimal MSC criteria, ALLOB® expressed significantly higher levels of chondro/osteoblastic genes such as BMP2 (fold change (FC) > 100), ALPL (FC > 12), CBFA1 (FC > 3) and differentiated significantly earlier than BM-MSC toward the osteogenic lineage. Moreover the bone formation model in nude mice demonstrated that used at the same cellular concentration, ALLOB® was able to induce significantly more (160% vs.107% for control animals) bone formation than BM-MSC (118% vs. 107 % for control animals) two weeks after administration. Conclusion: Our side-by-side comparison studies demonstrated that in vitro and in vivo, ALLOB® displays superior osteogenic capacity to BM-MScs and is therefore a better candidate for the treatment of bone fractures.

Keywords: gene expression, histomorphometry, mesenchymal stem cells, osteogenic differentiation potential, preclinical

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Authors: Yu-Mei Hsueh, Wei-Jen Chen, Yung-Kai Huang, Cheng-Shiuan Tsai, Kuo-Cheng Yeh

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Prostate cancer occurs in men over the age of 50, and rank sixth of the top ten cancers in Taiwan, and the incidence increased gradually over the past decade in Taiwan. Arsenic is confirmed as a carcinogen by International Agency for Research on (IARC). Arsenic induces oxidative stress may be a risk factor for prostate cancer, but the mechanism is not clear. Selenium is an important antioxidant element. Whether the association between plasma selenium levels and risk of prostate cancer are modified by different genotype of selenoprotein is still unknown. Glutathione peroxidase, selenoprotein P (SEPP1) and 15 kDa selenoprotein (SEP 15) are selenoprotein and regulates selenium transport and the oxidation and reduction reaction. However, the association between gene polymorphisms of selenoprotein and prostate cancer is not yet clear. The aim of this study is to determine the relationship between plasma selenium, polymorphism of selenoprotein, urinary total arsenic concentration and prostate cancer. This study is a hospital-based case-control study. Three hundred twenty-two cases of prostate cancer and age (±5 years) 1:1 matched 322 control group were recruited from National Taiwan University Hospital, Taipei Medical University Hospital, and Wan Fang Hospital. Well-trained personnel carried out standardized personal interviews based on a structured questionnaire. Information collected included demographic and socioeconomic characteristics, lifestyle and disease history. Blood and urine samples were also collected at the same time. The Research Ethics Committee of National Taiwan University Hospital, Taipei, Taiwan, approved the study. All patients provided informed consent forms before sample and data collection. Buffy coat was to extract DNA, and the polymerase chain reaction - restriction fragment length polymorphism (PCR-RFLP) was used to measure the genotypes of SEPP1 rs3797310, SEP15 rs5859, GPX1 rs1050450, GPX2 rs4902346, GPX3 rs4958872, and GPX4 rs2075710. Plasma concentrations of selenium were determined by inductively coupled plasma mass spectrometry (ICP-MS).Urinary arsenic species concentrations were measured by high-performance liquid chromatography links hydride generator and atomic absorption spectrometer (HPLC-HG-AAS). Subject with high education level compared to those with low educational level had a lower prostate cancer odds ratio (OR) Mainland Chinese and aboriginal people had a lower OR of prostate cancer compared to Fukien Taiwanese. After adjustment for age, educational level, subjects with GPX1 rs1050450 CT and TT genotype compared to the CC genotype have lower, OR of prostate cancer, the OR and 95% confidence interval (Cl) was 0.53 (0.31-0.90). SEPP1 rs3797310 CT+TT genotype compared to those with CC genotype had a marginally significantly lower OR of PC. The low levels of plasma selenium and the high urinary total arsenic concentrations had the high OR of prostate cancer in a significant dose-response manner, and SEPP1 rs3797310 genotype modified this joint association.

Keywords: prostate cancer, plasma selenium concentration, urinary total arsenic concentrations, glutathione peroxidase, selenoprotein P, selenoprotein 15, gene polymorphism

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Authors: Umara Nissar Rafiqi, Irum Gul, Nazima Nasrullah, Monica Saifi, Malik Z. Abdin

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Background: Stevia rebaudiana, a member of Asteraceae family is an important medicinal plant and produces a commercially used non-caloric natural sweetener, which is also an alternate herbal cure for diabetes. Steviol glycosides are the main sweetening compounds present in these plants. Secondary metabolites are crucial to the adaption of plants to the environment and its overcoming stress conditions. In agricultural procedures, the abiotic stresses like salinity, high metal toxicity and drought, in particular, are responsible for the majority of the reduction that differentiates yield potential from harvestable yield. Salt stress and heavy metal toxicity lead to increased production of reactive oxygen species (ROS). To avoid oxidative damage due to ROS and osmotic stress, plants have a system of anti-oxidant enzymes along with several stress induced enzymes. This helps in scavenging the ROS and relieve the osmotic stress in different cell compartments. However, whether stress induced toxicity modulates the activity of these enzymes in Stevia rebaudiana is poorly understood. Aim: The present study focussed on the effect of salinity, heavy metal toxicity (lead and mercury) on physiological traits and transcriptional profiling of Stevia rebaudiana. Method: Stevia rebaudiana plants were collected from the Central Institute of Medicinal and Aromatic plants (CIMAP), Patnagar, India and maintained under controlled conditions in a greenhouse at Hamdard University, Delhi, India. The plants were subjected to different concentrations of salt (0, 25, 50 and 75 mM respectively) and heavy metals, lead and mercury (0, 100, 200 and 300 µM respectively). The physiological traits such as shoot length, root numbers, leaf growth were evaluated. The samples were collected at different developmental stages and analysed for transcription profiling by RT-PCR. Transcriptional studies in stevia rebaudiana involves important antioxidant enzymes: catalase (CAT), superoxide dismutase (SOD), cytochrome P450 monooxygenase (CYP) and stress induced aquaporin (AQU), auxin repressed protein (ARP-1), Ndhc gene. The data was analysed using GraphPad Prism and expressed as mean ± SD. Result: Low salinity and lower metal toxicity did not affect the fresh weight of the plant. However, this was substantially decreased by 55% at high salinity and heavy metal treatment. With increasing salinity and heavy metal toxicity, the values of all studied physiological traits were significantly decreased. Chlorosis in treated plants was also observed which could be due to changes in Fe:Zn ratio. At low concentrations (upto 25 mM) of NaCl and heavy metals, we did not observe any significant difference in the gene expressions of treated plants compared to control plants. Interestingly, at high salt concentration and high metal toxicity, a significant increase in the expression profile of stress induced genes was observed in treated plants compared to control (p < 0.005). Conclusion: Stevia rebaudiana is tolerant to lower salt and heavy metal concentration. This study also suggests that with the increase in concentrations of salt and heavy metals, harvest yield of S. rebaudiana was hampered.

Keywords: Stevia rebaudiana, natural sweetener, salinity, heavy metal toxicity

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Authors: David Pinzauti, Simon De Jaegher, Maria D'Aguano, Manuele Biazzo

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The COVID-19 pandemic has left an indelible mark on global health, leading to a pressing need for understanding the intricate interactions between the virus and the human microbiome. This study focuses on characterizing the nasal microbiota of patients affected by COVID-19, with a specific emphasis on the comparison with unaffected individuals, to shed light on the crucial role of the microbiome in the development of this viral disease. To achieve this objective, Nanopore technology was employed to analyze the bacterial 16s rRNA full-length gene present in nasal swabs collected in Malta between January 2021 and August 2022. A comprehensive dataset consisting of 268 samples (126 SARS-negative samples and 142 SARS-positive samples) was subjected to a comparative analysis using an in-house, custom pipeline. The findings from this study revealed that individuals affected by COVID-19 possess a nasal microbiota that is significantly less diverse, as evidenced by lower α diversity, and is characterized by distinct microbial communities compared to unaffected individuals. The beta diversity analyses were carried out at different taxonomic resolutions. At the phylum level, Bacteroidota was found to be more prevalent in SARS-negative samples, suggesting a potential decrease during the course of viral infection. At the species level, the identification of several specific biomarkers further underscores the critical role of the nasal microbiota in COVID-19 pathogenesis. Notably, species such as Finegoldia magna, Moraxella catarrhalis, and others exhibited relative abundance in SARS-positive samples, potentially serving as significant indicators of the disease. This study presents valuable insights into the relationship between COVID-19 and the nasal microbiota. The identification of distinct microbial communities and potential biomarkers associated with the disease offers promising avenues for further research and therapeutic interventions aimed at enhancing public health outcomes in the context of COVID-19.

Keywords: COVID-19, nasal microbiota, nanopore technology, 16s rRNA gene, biomarkers

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Authors: Seyedeh Banafsheh Bagheri Marzouni, Sona Rostampour Yasouri

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Salmonella is one of the most important common pathogens between humans and animals worldwide. Globally, the prevalence of the disease in humans is due to the consumption of food contaminated with animal-derived Salmonella. These foods include eggs, red meat, chicken, and milk. Contamination of chicken and its products with Salmonella may occur at any stage of the chicken processing chain. Salmonella infection is usually not fatal. However, its occurrence is considered dangerous in some individuals, such as infants, children, the elderly, pregnant women, or individuals with weakened immune systems. If Salmonella infection enters the bloodstream, the possibility of contamination of tissues throughout the body will arise. Therefore, determining the potential risk of Salmonella at various stages is essential from the perspective of consumers and public health. The aim of this study is to isolate and identify Salmonella from chicken samples distributed in the Tehran market using the Gold standard culture method and PCR techniques based on specific genes, invA and ent. During the years 2022-2023, sampling was performed using swabs from the liver and intestinal contents of distributed chickens in the Tehran province, with a total of 120 samples taken under aseptic conditions. The samples were initially enriched in buffered peptone water (BPW) for pre-enrichment overnight. Then, the samples were incubated in selective enrichment media, including TT broth and RVS medium, at temperatures of 37°C and 42°C, respectively, for 18 to 24 hours. Organisms that grew in the liquid medium and produced turbidity were transferred to selective media (XLD and BGA) and incubated overnight at 37°C for isolation. Suspicious Salmonella colonies were selected for DNA extraction, and PCR technique was performed using specific primers that targeted the invA and ent genes in Salmonella. The results indicated that 94 samples were Salmonella using the PCR technique. Of these, 71 samples were positive based on the invA gene, and 23 samples were positive based on the ent gene. Although the culture technique is the Gold standard, PCR is a faster and more accurate method. Rapid detection through PCR can enable the identification of Salmonella contamination in food items and the implementation of necessary measures for disease control and prevention.

Keywords: culture, PCR, salmonella spp, salmonella enteritidis

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Authors: S. Woźniak, J. Rybka, T. Paszkowski, P. Milart

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A 30-year-old patient, who was pregnant for her second 9 weeks, was admitted to the hospital due to a suspected incomplete miscarriage. A fetal egg was found in the uterine cavity near the mouth of the fallopian tube. The patient was qualified for dilatation and curettage. The histopathological examination revealed fragments of the trophoblast. Two months later, the patient was re-admitted to the hospital due to vaginal bleeding and elevated levels of beta-hCG. Additional tests were performed. An intramural pregnancy was suspected. The patient was qualified for embolization of the uterine arteries and then treatment with methotrexate. Three weeks later, during a routine gynecological examination, a detached tumor 4 cm in diameter was found in the vagina. The material was sent for histopathological examination, which showed the presence of trophoblastic cells.

Keywords: ectopic pregnancy, intramural pregnancy, uterine artery embolization, methotrexate

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Authors: Aleksandra Bylinska, Samantha Slight-Webb, Kevin Thomas, Miles Smith, Susan Macwana, Nicolas Dominguez, Eliza Chakravarty, Joan T. Merrill, Judith A. James, Joel M. Guthridge

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Background: Systemic Lupus Erythematosus (SLE) is an interferon-related autoimmune disease characterized by B cell dysfunction. One of the main hallmarks is a loss of tolerance to self-antigens leading to increased levels of autoantibodies against nuclear components (ANAs). However, up to 20% of healthy ANA+ individuals will not develop clinical illness. SLE is more prevalent among women and minority populations (African, Asian American and Hispanics). Moreover, African Americans have a stronger interferon (IFN) signature and develop more severe symptoms. The exact mechanisms involved in ethnicity-dependent B cell dysregulation and the progression of autoimmune disease from ANA+ healthy individuals to clinical disease remains unclear. Methods: Peripheral blood mononuclear cells (PBMCs) from African (AA) and European American (EA) ANA- (n=12), ANA+ (n=12) and SLE (n=12) individuals were assessed by multimodal scRNA-Seq/CITE-Seq methods to examine differential gene signatures in specific B cell subsets. Library preparation was done with a 10X Genomics Chromium according to established protocols and sequenced on Illumina NextSeq. The data were further analyzed for distinct cluster identification and differential gene signatures in the Seurat package in R and pathways analysis was performed using Ingenuity Pathways Analysis (IPA). Results: Comparing all subjects, 14 distinct B cell clusters were identified using a community detection algorithm and visualized with Uniform Manifold Approximation Projection (UMAP). The proportion of each of those clusters varied by disease status and ethnicity. Transitional B cells trended higher in ANA+ healthy individuals, especially in AA. Ribonucleoprotein high population (HNRNPH1 elevated, heterogeneous nuclear ribonucleoprotein, RNP-Hi) of proliferating Naïve B cells were more prevalent in SLE patients, specifically in EA. Interferon-induced protein high population (IFIT-Hi) of Naive B cells are increased in EA ANA- individuals. The proportion of memory B cells and plasma cells clusters tend to be expanded in SLE patients. As anticipated, we observed a higher signature of cytokine-related pathways, especially interferon, in SLE individuals. Pathway analysis among AA individuals revealed an NRF2-mediated Oxidative Stress response signature in the transitional B cell cluster, not seen in EA individuals. TNFR1/2 and Sirtuin Signaling pathway genes were higher in AA IFIT-Hi Naive B cells, whereas they were not detected in EA individuals. Interferon signaling was observed in B cells in both ethnicities. Oxidative phosphorylation was found in age-related B cells (ABCs) for both ethnicities, whereas Death Receptor Signaling was found only in EA patients in these cells. Interferon-related transcription factors were elevated in ABCs and IFIT-Hi Naive B cells in SLE subjects of both ethnicities. Conclusions: ANA+ healthy individuals have altered gene expression pathways in B cells that might drive apoptosis and subsequent clinical autoimmune pathogenesis. Increases in certain regulatory pathways may delay progression to SLE. Further, AA individuals have more elevated activation pathways that may make them more susceptible to SLE.

Keywords:

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Authors: Vinai K. Singh

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In Neural Network-based Learning techniques, there are several models of Convolutional Networks. Whenever the methods are deployed with large datasets, only then can their applicability and appropriateness be determined. Clinical and pathological pictures of lobular carcinoma are thought to exhibit a large number of random formations and textures. Working with such pictures is a difficult problem in machine learning. Focusing on wet laboratories and following the outcomes, numerous studies have been published with fresh commentaries in the investigation. In this research, we provide a framework that can operate effectively on raw photos of various resolutions while easing the issues caused by the existence of patterns and texturing. The suggested approach produces very good findings that may be used to make decisions in the diagnosis of cancer.

Keywords: lobular carcinoma, convolutional neural networks (CNN), deep learning, histopathological imagery scans

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Authors: Mei Chen, Yingping Hou, Michelle Hao, Soheil Aghamohammadzadeh, Esteban Terzo, Roger Clark, Vijay Modur

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Background: Recessive Dystrophic Epidermolysis Bullosa (RDEB) is a genetic skin condition characterized by skin tearing and unremitting blistering upon minimal trauma. Repeated blistering, fibrosis, and scarring lead to aggressive squamous cell carcinoma later in life. RDEB is caused by mutations in the COL7A1 gene encoding collagen type VII (C7), the major component of anchoring fibrils mediating epidermis-dermis adherence. Nonsense mutations in the COL7A1 gene of a subset of RDEB patients leads to premature termination codons (PTC). Similarly, most Junctional Epidermolysis Bullosa (JEB) cases are caused by nonsense mutations in the LAMB3 gene encoding the β3 subunit of laminin 332. Currently, there is an unmet need for the treatment of RDEB and JEB. Zikani Therapeutics has discovered an array of macrocyclic compounds with ring structures similar to macrolide antibiotics that can facilitate readthrough activity of nonsense mutations in the COL7A1 and LAMB3 genes by acting as Ribosome Modulating Agents (RMAs). The medicinal chemistry synthetic advancements of these macrocyclic compounds have allowed targeting the human ribosome while preserving the structural elements responsible for the safety and pharmacokinetic profile of clinically used macrolide antibiotics. Methods: C7 expression was used as a measure of readthrough activity by immunoblot assays in two primary human fibroblasts from RDEB patients (R578X/R578X and R163X/R1683X-COL7A1). Similarly, immunoblot assays in C325X/c.629-12T > A-LAMB3 keratinocytes were used to measure readthrough activity for JEB. The relative readthrough activity of each compound was measured relative to Gentamicin. An imaging-based fibroblast migration assay was used as an assessment of C7 functionality in RDEB-fibroblasts over 16-20 hrs. The incubation period for the above experiments was 48 hrs for RDEB fibroblasts and 72 hours for JEB keratinocytes. Results: 9 RMAs demonstrated increased protein expression in both patient RDEB fibroblasts. The highest readthrough activity at tested concentrations without cytotoxicities increased protein expression up to 179% of Gentamicin (400 µg/ml), with favored readthrough activity in R163X/R1683X-COL7A1 fibroblasts. Concurrent with protein expression, fibroblast hypermotility phenotype observed in RDEB was rescued by reducing motility by ~35% to WT levels (the same level as 690 µM Gentamicin treated cells). Laminin β3 expression was also shown to be increased by 6 RMAs in keratinocytes to 33-83% of (400 µg/ml) Gentamicin. Conclusions: To date, 9 RMAs have been identified that enhance the expression of functional C7 in a mutation-dependent manner in two different RDEB patient fibroblast backgrounds (R578X/R578X and R163X/R1683X-COL7A1). A further 6 RMAs have been identified that enhance the readthrough of C325X-LAMB3 in JEB patient keratinocytes. Based on the clinical trial conducted by us with topical gentamycin in 2017, Zikani’s RMAs achieve clinically significant levels of read-through for the treatment of recessive dystrophic and Junctional Epidermolysis Bullosa.

Keywords: epidermolysis bullosa, nonsense mutation, readthrough, ribosome modulation

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Authors: Natalia Moreno-Castellanos, Amaia Rodriguez, Gema Frühbeck

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Introduction: In adipocytes, the cytoskeleton undergoes important expression and distribution in adipocytes rearrangements during adipogenesis and in obesity. Indeed, a role for these proteins in the regulation of adipocyte differentiation and response to insulin has been demonstrated. Recently, septins have been considered as new components of the cytoskeletal network that interact with other cytoskeletal elements (actin and tubulin) profoundly modifying their dynamics. However, these proteins have not been characterized as yet in adipose tissue. In this work, were examined the cellular, molecular and functional features of a member of this family, septin 11 (SEPT11), in adipocytes and evaluated the impact of obesity on the expression of this protein in human adipose tissue. Methods: Adipose gene and protein expression levels of SEPT11 were analysed in human samples. SEPT11 distribution was evaluated by immunocytochemistry, electronic microscopy, and subcellular fractionation techniques. GST-pull down, immunoprecipitation and a Yeast-Two Hybrid (Y2H) screening were used to identify the SEPT11 interactome. Gene silencing was employed to assess the role of SEPT11 in the regulation of insulin signaling and lipid metabolism in adipocytes. Results: SEPT11 is expressed in human adipocytes, and its levels increased in both omental and subcutaneous adipose tissue in obesity, with SEPT11 mRNA content positively correlating with parameters of insulin resistance in subcutaneous fat. In non-stimulated adipocytes, SEPT11 immunoreactivity showed a ring-like distribution at the cell surface and associated to caveolae. Biochemical analyses showed that SEPT11 interacted with the main component of caveolae, caveolin-1 (CAV1) as well as with the fatty acid-binding protein, FABP5. Notably, the three proteins redistributed and co-localized at the surface of lipid droplets upon exposure of adipocytes to oleate. In this line, SEPT11 silencing in 3T3-L1 adipocytes impaired insulin signaling and decreased insulin-induced lipogenesis. Conclusions: Those findings demonstrate that SEPT11 is a novel component of the adipocyte cytoskeleton that plays an important role in the regulation of lipid traffic, metabolism and can thus represent a potential biomarker of insulin resistance in obesity in adipocytes through its interaction with both CAV1 and FABP5.

Keywords: caveolae, lipid metabolism, obesity, septins

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Authors: Karthikeyan M., Paul M. J.

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This abstract describes a case of central pancreatectomy (CP) for a 50-year-old woman with a neuroendocrine tumor in the mid-body of the pancreas. CP, a parenchyma-sparing surgical option, preserves the distal pancreas and spleen, reducing the risk of pancreatic endocrine and exocrine insufficiency compared to traditional resections. The patient, initially misdiagnosed with transient ischemic attack, presented with hypoglycemic symptoms and was found to have a pancreatic lesion. Post-operative results were positive, with a reduction in pancreatic drain volume and normalization of blood sugar levels. This case highlights CP's efficacy in treating centrally located pancreatic lesions while maintaining pancreatic function.

Keywords: central pancreatectomy without anastomosis, no endocrine deficiency on follow-op, less post-op hospital stay, less post-op complications

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Authors: Sandeep Mohindra

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Since 2013, the authors have operated on 568 cases of pituitary tumors through the trans-sphenoidal route, using the binostril approach. The distribution included 486 cases of non-functioning pituitary tumors, 24 cases of Growth hormone(GH) secreting tumors(acromegaly), and 28 cases of adrenocorticotrophic(ACTH) secreting tumors(Cushing's Disease). The authors utilized neuro-navigation for 18 cases, and all belonged to the functional tumor category. Complications included ICA injury in 2 cases, fatal meningitis in 5 cases, while CSF leak required repair in 28 cases. Satisfactory excision was noted in 512 cases, while recurrence/residual required repeat surgery in 32 cases. Authors conclude that trans sphenoidal route remains the best and optimal way of managing sellar tumors, especially pituitary adenomas.

Keywords: pituitary, adenoma, trans-sphenoidal, endonasal, neuronavigation

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Authors: Asma Rashid, Shazia Iram, Iftikhar Ahmad

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Mango sudden death is an emerging problem in Pakistan. As its prevalence is observed in almost all mango growing areas and severity varied from 2-5% in Punjab and 5-10% in Sindh. Symptoms on affected trees include bark splitting, discoloration of the vascular tissue, wilting, gummosis and at the end rapid death. Total of n= 45 isolates were isolated from different mango growing areas of Punjab and Sindh. Pathogenicity of these fungal isolates was tested through artificial inoculation method on different hosts (potato tubers, detached mango leaves, detached mango twigs and mango plants) under controlled conditions and all were proved pathogenic with varying degree of aggressiveness in reference to control. The findings of the present study proved that out of these four methods, potato tubers inoculation method was the most ideal as this fix the inoculums on the target site. Increased fungal growth and spore numbers may be due to soft tissues of potato tubers from which Ceratocystis isolates can easily pass. Lesion area on potato tubers was in the range of 7.09-0.14 cm2 followed by detached mango twigs which were ranged from 0.48-0.09 cm2). All pathological results were proved highly significant at P<0.05 through ANOVA but isolate to isolate showed non-significant behaviour but they have the positive effect on lesion area. Re-isolation of respective fungi was achieved with 100 percent success which results in the verification of Koch’s postulates. DNA of fungal pathogens was successfully extracted through phenol chloroform method. Amplification was done through ITS, b-tubulin gene, and Transcription Elongation Factor (EF1-a) gene primers and the amplified amplicons were sequenced and compared from NCBI which showed 99-100 % similarity with Ceratocystis manginecans. Fungus Ceratocystis manginecans formed one of strongly supported sub-clades through phylogenetic tree. Results obtained through this work would be supportive in establishment of relation of isolates with their region and will give information about pathogenicity level of isolates that would be useful to develop the management policies to reduce the afflictions in orchards caused by mango sudden death.

Keywords: artificial inoculation, mango, Ceratocystis manginecans, phylogenetic, screening

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Authors: Hosam Abdelhady

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Filming the behaviors of individual DNA molecules in their environment when they interact with individual medicinal nano-polymers in a molecular scale has opened the door to understand the effect of the molecular shape, size, and incubation time with nanocarriers on optimizing the design of robust genetic Nano molecules able to resist the enzymatic degradation, enter the cell, reach to the nucleus and kill individual cancer cells in their environment. To this end, we will show how we applied the 4D AFM as a guide to finetune the design of genetic nanoparticles and to film the effects of these nanoparticles on the nanomechanical and morphological profiles of individual cancer cells.

Keywords: AFM, dendrimers, nanoparticles, DNA, gene therapy, imaging

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Authors: Azza El-Medany, Jamila El-Medany

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Residential and agricultural pesticide use is widespread in the world. Their extensive and indiscriminative use, in addition with their ability to interact with biological systems other than their primary targets constitute a health hazards to both humans and animals. The toxic effects of pesticides include alterations in metabolism; there is a lack of knowledge that organophosphates can cause pancreatic toxicity. The primary goal of this work is to study the effects of chronic exposure to Diazinon an organophosphate used in agriculture on pancreatic tissues and evaluate the ameliorating effect of Mesna as antioxidant on the toxicity of Diazinon on pancreatic tissues.40 adult male rats, their weight ranged between 300-350 g. The rats were classified into three groups; control (10 rats) was received corn oil at a dose of 1 0 mg/kg/day by gavage once a day for 2 months. Diazinon (15 rats) was received Diazinon at a dose of 10 mg/kg/day dissolved in corn oil by gavage once a day for 2 months. Treated group (15 rats), were received Mesna 180mg/kg once a week by gavage 15 minutes before administration of Diazinon for 2 months. At the end of the experiment, animals were anesthetized, blood samples were taken by cardiac puncture for glucose and insulin assays and pancreas was removed and divided into 3 portions; first portion for histopathological study; second portion for ultrastructural study; third portion for biochemical study using Elisa Kits including determination of malondialdehyde (MDA), tumor necrosis factor α (TNF-α), myeloperoxidase activity (MPO), interleukin 1β (IL-1β). A significant increase in the levels of MDA, TNF-α, MPO activity, IL-1β, serum glucose levels in the toxicated group with Diazinon were observed, while a significant reduction was noticed in GSH in serum insulin levels. After treatment with Mesna a significant reduction was observed in the previously mentioned parameters except that there was a significant rise in GSH in insulin levels. Histopathological and ultra-structural studies showed destruction in pancreatic tissues and β cells were the most affected cells among the injured islets as compared with the control group. The current study try to spot light about the effects of chronic exposure to pesticides on vital organs as pancreas also the role of oxidative stress that may be induced by them in evoking their toxicity. This study shows the role of antioxidant drugs in ameliorating or preventing the toxicity. This appears to be a promising approach that may be considered as a complementary treatment of pesticide toxicity.

Keywords: Diazinon, reduced glutathione, myeloperoxidase activity, tumor necrosis factor α, Mesna

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