Search results for: reconstitution

Authors: Wei Wang, Martin Chen

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The reconstitution behaviours of nutritional products could impact user experience. Reconstitution issues such as lump formation and white flecks sticking to bottles surfaces could be very unappealing for the consumers in milk preparation. The controlling steps in dissolving instant milk powders include wetting, swelling, sinking, dispersing, and dissolution as in the literature. Each stage happens simultaneously with the others during milk preparation, and it is challenging to isolate and measure each step individually. This study characterized three adult nutritional products for different properties including particle size, density, dispersibility, stickiness, and capillary wetting to understand the relationship between powder physical properties and their reconstitution behaviours. From the results, the formation of clumps can be caused by different factors limiting the critical steps of powder reconstitution. It can be caused by small particle size distribution, light particle density limiting powder wetting, or the rapid swelling and dissolving of particle surface materials to impede water penetration in the capillary channels formed by powder agglomerates. For the grain or white flecks formation in milk preparation, it was believed to be controlled by dissolution speed of the particles after dispersion into water. By understanding those relationship between fundamental powder structure and their user experience in reconstitution, this information provides us new and multiple perspectives on how to improve the powder characteristics in the commercial manufacturing.

Keywords: characterization, dairy nutritional powder, physical property, reconstitution

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Authors: Mohammaad Afzal Khan, Ghazi Abdulmalik Ashoor , Fatimah Alanazi, Talal Shamma, Abdullah Altuhami, Hala Abdalrahman Ahmed, Abdullah Mohammed Assiri, Dieter Clemens Broering

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Microvascular injuries during inflammation are key causes of transplant malfunctioning and permanent failure, which play a major role in the development of chronic rejection of the transplanted organ. Inflammation-induced microvascular loss is a promising area to investigate the decisive roles of regulatory and effector responses. The present study was designed to investigate the impact of IL-10 on immunotolerance, in particular, the microenvironment of the allograft during rejection. Here, we investigated the effects of IL-10 blockade/ reconstitution and serially monitored regulatory T cells (Tregs), graft microvasculature, and airway epithelium in rejecting airway transplants. We demonstrated that the blocking/reconstitution of IL-10 significantly modulates CD4+FOXP3+ Tregs, microvasculature, and airway epithelium during rejection. Our findings further highlighted that blockade of IL-10 upregulated proinflammatory cytokines, IL-2, IL-1β, IFN-γ, IL-15, and IL-23, but suppressed IL-5 secretion during rejection; however, reconstitution of IL-10 significantly upregulated CD4+FOXP3+ Tregs, tissue oxygenation/blood flow and airway repair. Collectively, these findings demonstrate a potential reparative modulation of IL-10 during microvascular and epithelial repair, which could provide a vital therapeutic window to rejecting transplants in clinical practice.

Keywords: interleukin -10, regulatory T cells, allograft rejection, immunotolerance

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Authors: R. Burahee, B. Chassinat, T. de Laclos, A. Dépée, A. Sastim

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The purpose of this paper is to propose a solution to detect, predict and reconstitute rain drops in real time – during the night – using an embedded material with an infrared camera. To prevent the system from needing too high hardware resources, simple models are considered in a powerful image treatment algorithm reducing considerably calculation time in OpenCV software. Using a smart model – drops will be matched thanks to a process running through two consecutive pictures for implementing a sophisticated tracking system. With this system drops computed trajectory gives information for predicting their future location. Thanks to this technique, treatment part can be reduced. The hardware system composed by a Raspberry Pi is optimized to host efficiently this code for real time execution.

Keywords: reconstitution, prediction, detection, rain drop, real time, raspberry, infrared

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Authors: W. Abu Rayyan, A. Singh, A. M. Al-Jaafreh, W. Abu Dayyih, M. Bustami, S. Salem, N. Seder, K. Schröppel

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Hyphal growth and the transcriptional regulation to the host environment are key issues during the pathogenesis of C. albicans. Tec1p is the C. albicans homolog of a TEA transcription factor family, which share a conserved DNA-binding TEA domain in their N-terminal. In order to define a structure-function relationship of the C. albicans Tec1p protein, we constructed several mutations on the N terminal, C terminal or in the TEA binding domain itself by homologous recombination technology. The modifications in the open reading frame of TEC1 were tested for reconstitution of the morphogenetic development of the tec1/tec1 mutant strain CaAS12. Mutation in the TEA consensus sequence did not confer transition to hyphae whereas the reconstitution of the full-length Tec1p has reconstituted hyphal development. A deletion in one of glutamine-rich regions either in the Tec1p N-terminal or the C-terminal in regions of 53-212 or 637–744 aa, respectively, did not restore morphological development in mutant CaAS12 strain. Whereas, the reconstitution with Tec1p mutants other than the glutamate-rich region has restored the morphogenetic switch. Additionally, the deletion of the glutamine-rich region has attenuated the invasive growth and the heat shock resistance of C. albicans. In conclusion, we show that a glutamine-rich region of Tec1p is essential for the hyphal development and mediating adaptation to the host environment of C. albicans.

Keywords: Candida albicans, morphogenetic development, TEA domain, hyphal formation, TEC1

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Authors: Nadine Ali Hassan, Ngoc Son Nguyen, Didier Marot, Fateh Bendahmane

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This paper aims at presenting a study of the effect of scalping methods on the mechanical properties of coarse soils by resorting to numerical simulations based on the discrete element method (DEM) and experimental triaxial tests. Two reconstitution methods are used, designated as scalping method and substitution method. Triaxial compression tests are first simulated on a granular materials with a grap graded particle size distribution by using the DEM. We study the effect of these reconstitution methods on the stress-strain behavior of coarse soils with different fine contents and with different ways to control the densities of the scalped and substituted materials. Experimental triaxial tests are performed on original mixtures of sands and gravels with different fine contents and on their corresponding scalped and substituted samples. Numerical results are qualitatively compared to experimental ones. Agreements and discrepancies between these results are also discussed.

Keywords: coarse soils, mechanical behavior, scalping, replacement, triaxial devices

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Authors: S. Mohd Afzal, R. O'Connell

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Kaposi's sarcoma (KS) is the most common HIV-related cancer, and ocular manifestations constitute at least 25% of all KS cases. However, ocular presentations often occur in the context of systemic KS, and isolated lesions are rare. We report a unique case of ocular KS masquerading as subconjunctival haemorrhage, and only developing systemic manifestations after initiation of HIV treatment. Case: A 49-year old man with previous hypertensive stroke and newly diagnosed HIV infection presented with an acutely red left eye following repeated bouts of coughing. Given the convincing history of poorly controlled hypertension and cough, a diagnosis of subconjunctival haemorrhage was made. Over the next week, his ocular lesion began to improve and he subsequently started anti-retroviral therapy. Prior to receiving anti-retroviral therapy, his CD4+ lymphocyte count was 194 cells/mm3 with HIV viral load greater than 1 million/ml. This rapidly improved to a viral load of 150 copies/ml within 2 weeks of starting treatment. However, a few days after starting HIV treatment, his ocular lesion recurred. Ophthalmic examination was otherwise normal. He also developed widespread lymphadenopathy and multiple dark lesions on his torso. Histology and virology confirmed KS, systemically triggered by Immune Reconstitution Syndrome (KS-IRIS). The patient has since undergone chemotherapy successfully. Discussion: Kaposi's sarcoma is an atypical tumour caused by human herpesvirus 8 (HHV-8), also known as Kaposi’s sarcoma-associated herpesvirus (KSHV). In immunosuppressed patients, KSHV can also cause lymphoproliferative disorders such as primary effusion lymphoma and Castleman's disease (in our patient’s case, this was excluded through histological analysis of lymph nodes). KSHV is one of the seven currently known human oncoviruses, and its pathogenesis is poorly understood. Up to 13% of patients with HIV-related KS experience worsening of the disease after starting anti-retroviral treatment, due to a sudden increase in CD4 cell counts. Histology remains the diagnostic gold standard. Current British HIV Association (BHIVA) guidelines recommend treatment using anti-retroviral drugs, with either intralesional vinblastine for local disease or systemic chemotherapy for disseminated KS. Conclusion: This case is unique as ocular KS as initial presentation is rare and our patient's diagnosis was only made after systemic lesions were triggered by immune reconstitution. KS should be considered as an important differential diagnosis for red eyes in all patients at risk of acquiring HIV infection.

Keywords: human herpesvirus 8, human immunodeficiency virus, immune reconstitution syndrome, Kaposi’s sarcoma, Kaposi’s sarcoma-associated herpesvirus

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Authors: Mbaeyi-Nwaoha, Ifeoma Elizabeth, Orngu, Africa Orngu

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Composite flours were processed from blends of yellow maize (Zea mays), sesame seed (Sesamum indicum) and oyster mushroom (Pleurotus ostreatus) powder in the ratio of 80:20:0; 75:20:5; 70:20:10; 65:20:10 and 60:20:20, respectively to produce the breakfast cereal coded as YSB, SMB, TMB, PMB and OMB with YSB as the control. The breakfast cereals were produced by hydration and toasting of yellow maize and sesame to 160oC for 25 minutes and blended together with oven dried and packaged oyster mushroom. The developed products (flours and breakfast cereals) were analyzed for proximate composition, vitamins, minerals, anti-nutrients, phytochemicals, functional, microbial and sensory properties. Results for the flours showed: proximate composition (%): moisture (2.59-7.27), ash (1.29-7.57), crude fat (0.98-14.91), fibre (1.03-16.02), protein (10.13-35.29), carbohydrate (75.48-38.18) and energy (295.18-410.75kcal). Vitamins ranged as: vitamin A (0.14-9.03 ug/100g), vitamin B1 (0.14-0.38), vitamin B2 (0.07-0.15), vitamin B3(0.89-4.88) and Vitamin C (0.03-4.24). Minerals (mg/100g) were reported thus: calcium (8.01-372.02), potassium (1.40-1.85), magnesium (12.09-13.15), iron (1.23-5.25) and zinc (0.85-2.20). The results for anti-nutrients and phytochemical ranged from: tannin (1.50-1.61mg/g), Phytate (0.40-0.71mg/g), Oxalate(1.81-2.02mg/g), Flavonoid (0.21-1.27%) and phenolic (1.12-2.01%). Functional properties showed: bulk density (0.51-0.77g/ml), water absorption capacity (266.0-301.5%), swelling capacity (136.0-354.0%), least Gelation (0.55-1.45g/g) and reconstitution index (35.20-69.60%). The total viable count ranged from 6.4× 102to1.0× 103cfu/g while the total mold count was from 1.0× 10to 3.0× 10 cfu/g. For the breakfast cereals, proximate composition (%) ranged thus: moisture (4.07-7.08), ash (3.09-2.28), crude fat(16.04-12.83), crude fibre(4.30-8.22), protein(16.14-22.54), carbohydrate(56.34-47.04) and energy (434.34-393.83Kcal).Vitamin A (7.99-5.98 ug/100g), vitamin B1(0.08-0.42mg/100g), vitamin B2(0.06-0.15 mg/100g), vitamin B3(1.91-4.52 mg/100g) and Vitamin C(3.55-3.32 mg/100g) were reported while Minerals (mg/100g) were: calcium (75.31-58.02), potassium (0.65-4.01), magnesium(12.25-12.62), iron (1.21-4.15) and zinc (0.40-1.32). The anti-nutrients and phytochemical revealed the range (mg/g) as: tannin (1.12-1.21), phytate (0.69-0.53), oxalate (1.21-0.43), flavonoid (0.23-1.22%) and phenolic (0.23-1.23%). The bulk density (0.77-0.63g/ml), water absorption capacity (156.5-126.0%), swelling capacity (309.5-249.5%), least gelation (1.10-0.75g/g) and reconstitution index (49.95-39.95%) were recorded. From the total viable count, it ranged from 3.3× 102to4.2× 102cfu/g but no mold growth was detected. Sensory scores revealed that the breakfast cereals were acceptable to the panelist with oyster mushroom supplementation up to 10%.

Keywords: oyster mushroom (Pleurotus ostreatus), sesame seed (Sesamum indicum), yellow maize (Zea mays, instant breakfast cereals

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Authors: Vijayalakshmi Marella, NageswaraRaoPilli

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This paper describes a simple, rapid and sensitive liquid chromatography / tandem mass spectrometry assay for the determination of montelukast in human plasma using montelukast d6 as an internal standard. Analyte and the internal standard were extracted from 50 µL of human plasma via solid phase extraction technique without evaporation, drying and reconstitution steps. The chromatographic separation was achieved on a C18 column by using a mixture of methanol and 5mM ammonium acetate (80:20, v/v) as the mobile phase at a flow rate of 0.8 mL/min. Good linearity results were obtained during the entire course of validation. Method validation was performed as per FDA guidelines and the results met the acceptance criteria. A run time of 2.5 min for each sample made it possible to analyze more number of samples in short time, thus increasing the productivity. The proposed method was found to be applicable to clinical studies.

Keywords: Montelukast, tandem mass spectrometry, montelukast d6, FDA guidelines

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Authors: Kanchana Sitlaothaworn

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Yogurt capsule was made by mixing 14% w/v of reconstitution of skim milk with 2% FOS. The mixture was fermented by commercial yogurt starter comprising Lactobacillus bulgaricus and Streptococcus thermophilus. These yogurts were made as yogurt powder by freeze-dried. Yogurt powder was put into capsule then stored for 28 days at 4oc. 8ml of commercial yogurt was found to be the most suitable inoculum size in yogurt production. After freeze-dried, the viability of L. bulgaricus and S. thermophilus reduced from 109 to 107 cfu/g. The precence of sucrose cannot help to protect cell from ice crystal formation in freeze-dried process, high (20%) sucrose reduced L. bulgaricus and S. thermophilus growth during fermentation of yogurt. The addition of FOS had reduced slowly the viability of both L. bulgaricus and S. thermophilus similar to control (without FOS) during 28 days of capsule storage. The viable cell exhibited satisfactory viability level in capsule storage (6.7x106cfu/g) during 21 days at 4oC.

Keywords: yogurt capsule, Lactobacillus bulgaricus, Streptococcus thermophilus, freeze-drying, sucrose

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Authors: Mehaiguene Madjid, Touhari Fadhila, Meddi Mohamed

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The Cheliff basin offers a good hydrological example for the possibility of studying the problem which elucidated in the future, because of the unclearity in several aspects and hydraulic installation. Thus, our study of the Cheliff basin is divided into two principal parts: The spatial evaluation of the precipitation: also, the understanding of the modes of the reconstitution of the resource in water supposes a good knowledge of the structuring of the precipitation fields in the studied space. In the goal of a good knowledge of revitalizes them in water and their management integrated one judged necessary to establish a precipitation card of the Cheliff basin for a good understanding of the evolution of the resource in water in the basin and that goes will serve as basis for all study of hydraulic planning in the Cheliff basin. Then, the establishment of the precipitation card of the Cheliff basin answered a direct need of setting to the disposition of the researchers for the region and a document of reference that will be completed therefore and actualized. The hydrological study, based on the statistical hydrometric data processing will lead us to specify the hydrological terms of the assessment hydrological and to clarify the fundamental aspects of the annual flow, seasonal, extreme and thus of their variability and resources surface water.

Keywords: hydrological assessment, surface water resources, Cheliff, Algeria

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Authors: Susmita Roy, Madhavan Nallani

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In recent years' polymersomes, self-assembled polymeric vesicles emerge from block copolymers, have been widely investigated due to their enhance stability and unique advantageous properties compared to their phospholipid counterpart, liposomes, dendrimers, and micelles. It provides a distinctive platform for advanced therapeutics and the creation of complex (bio) catalytically active systems for research in Nanomedicine and synthetic biology. Inspired by nature, where compartmentalization of biological components is all ubiquitous, we are interested in developing a platform technology of self-assembled multifunctional compartments with applications in areas from targeted drug/gene delivery, biosensing, pharmaceutical to cosmetics. Polymersome surfaces can be a proper choice of derivatization with a controlled amount of functional groups. To achieve site-specific targeting of polymersomes, biological recognition motives can be attached to the polymersomes surface by standard bioconjugation techniques, (like esterification, amidation, thiol-maleimide coupling, click-chemistry routes or other coupling methods). Herein, we are developing easy going, one-step bioconjugation strategies for site-specific surface functionalized biodegradable polymeric and/or polymer-lipid hybrid vesicles for targeted drug delivery. Biodegradable polymer, polycaprolactone-b-polyethylene glycol (PCL-PEG), polylactic acid-b-polyethylene glycol (PLA-PEG) and phospholipid, 1-palmitoyl-2- oleoyl-sn-glycero-3-phosphocholine (POPC) has been widely used for numerous vesicle formulations. Some of these drug-loaded formulations are being tested on mice for controlled release. These surface functionalized polymersomes are also appropriate for membrane protein reconstitution/insertion, antibodies conjugation and various bioconjugation with diverse targeted molecules for controlled drug delivery.

Keywords: drug delivery, membrane protein, polymersome, surface modification

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Authors: Olga Shuvaeva, Tamara Romanova, Sergey Volynkin, Valentina Podolinnaya

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Green chemistry concept is focused on the prevention of environmental pollution caused by human activity. However, there are a lot of contaminated areas in the world which pose a serious threat to ecosystems in terms of their conservation. Therefore in accordance with the principles of green chemistry, it should not be forgotten about the need to clean these areas. Furthermore, the waste material often contains the valuable components, the extraction of which by traditional wet chemical technologies is inefficient both from the economic and environmental protection standpoint. Wherein, the plants may be successfully used to ‘scavenge’ a range of metals from polluted land sites in an approach allowing to carry out both of these processes – phytoremediation and phytomining in conjunction. The goal of the present work was to study bioaccumulation ability of floating macrophytes such as water hyacinth and pondweed toward Hg, Ba, Cd, Mo and Pb as pollutants in aquatic medium and terrestrial plants (birch, reed, and cane) towards gold and silver as valuable components. The peculiarity of ongoing research was that the plants grew under extreme conditions (pH of drainage and pore waters was about 2.5). The study was conducted at the territory of Ursk tailings (Southwestern Siberia, Russia) formed as a result of primary polymetallic ores cyanidation. The waste material is mainly presented (~80%) by pyrite (FeS₂) and barite (BaSO₄), the raw minerals included FeAsS, HgS, PbS, Ag₂S as minor ones. It has been shown that water hyacinth demonstrates high ability to accumulate different metals, and what is especially important – to remove mercury from polluted waters with BCF value more than 1000. As for the gold, its concentrations in reed and cane growing near the waste material were estimated as 500 and 900 μg∙kg⁻¹ respectively. It was also found that the plants can survive under extreme conditions of acidic environment and hence we can assume that there is a principal opportunity to use them for the valuable substances extraction from an area of the mining waste dumps burial.

Keywords: bioaccumulation, gold, heavy metals, mine tailing

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Authors: Toumi Farah, Charallah Salima

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In the Algerian Sahara, goat farming is predominant, and it’s associated with other types of breeding, particularly camel and sheep; it also constitutes a significant proportion of breeding exclusively goat. This Saharan goat is a small ruminant with a black dress with white’s spots, hanging ears, and a coat more or less long. It is known for its hardiness and resistance to adverse conditions of arid zones and its perfect ecophysiological adaptation to harsh environmental conditions. However, pregnancy alterations, particularly abortion, degrade its productivity and cause economic losses, having both direct and indirect effects on animal production, like the costs of veterinary interventions and the reconstitution of livestock. The purpose of this work is to study the histological aspect of the liver of goats’ aborted living under nomadic herds in the region of Béni-Abbès (30° 7' N, 2° 10 'O). The organs were collected in physiological serum, rinsed, and then fixed with formaldehyde (37°, diluted at 10%). After that, these samples were processed for a topographic study. The morphometric study of the liver was performed by using an image analysis and processing software "Image J"; the various measurements obtained are intended to specify the supposed stage of development according to the body weight. The histological structure of the liver shows that the hepatic parenchyma consists of vascular conjunctive spaces surrounded by Glisson’s capsule. The sinusoids and hepatic portal vein are full of red blood cells, representing sinusoidal congestion and a thrombosed vein. At high magnification, the blood vessels show the presence of vascular thrombosis and haemorrhage in some areas of the hepatic parenchyma. Morphometric analysis shows that the number of liver parenchymal cells and the diameter of liver vessels vary according to the stage of development. The results obtained will provide details of the anatomical and cellular elements that can be used in the diagnosis of early or late abortion and late embryonic death. It would be interesting to find, by immunohistochemistry, some inflammatory markers useful for monitoring the progress of pregnancy and bioindicators of fetomaternal distress.

Keywords: aborting goat, arid zone, liver, histopathology

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Authors: Dawa Wangmo

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The anti-religious campaigns of the Soviet Union in the 1930s eradicated Kalmyk Buddhism from the public sphere. Following Perestroika, the Kalmyks retained a sense of being essentially Buddhist people. Nevertheless, since the collapse of the Soviet Communist regime, Kalmykia has been going through vigorous ethnic and cultural revitalization. The new Kalmyk government is reviving the religion with the building of Buddhist temples and the attempted training of Kalymk monks. Kalmykia, officially an autonomous republic within the Federation of Russia, is situated in the European part of Russia in the steppe region bordering the Caspian Sea in its southeast. According to the 2010 census, the Kalmyks, a people of Mongolian origin, constitute over 57 percent of the Republic’s population of less than 290000. Russians living in Kalmykia comprise around 30 percent, the remainder being various Slavic and Asian groups. Since the Kalmyks historically adhere to Buddhism, Kalmykia is often described in tourist brochures and proudly by the Kalmyks themselves as one of the three “traditional Buddhist republics” of Russia and “the only Buddhist region” in Europe. According to traditional Kalmyk Gelug Buddhism, monasticism is the central aspect; hence monastic Tibetans from India have been invited to the Republic to help revive Buddhism and their Buddhist identity in Russia as a whole. However, for the young post-soviets, the monastic way of life is proving too alien, and the subsequent labeling by these monks of ‘surviving’ Kalmyk Buddhist practices as superstitious, mistaken, or corrupt is an initial step in the purification of alternate views, leading to religious reform. This sentiment is also felt by younger Kalmyks who do not find sense in surviving Buddhism but believe more in the philosophical approach of Buddhism taught by the visiting Buddhist teachers at Dharma centers. By discussing this post-soviet shift in local notions of religious efficacy, an attempt will be made to shed light on how the social movements of both reform and revival arise as a collusion between contemporary Tibetan and Kalmyk views on the nature of true Buddhism. This work explores aspects of religious innovation that have developed since the early 1990s in the process of reconstitution of ethnic and religious identity in Kalmykia, a Republic in the southwest of Russia. Any attempts to study the history of Buddhism in Kalmykia would surely mean studying the “History of the most northern Dharma community in the World.”

Keywords: Kalmykia, Tibetan Buddhism, reform, revival, identity

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Authors: Lea Boidin, Martha Baydoun, Bertrand Leroux, Olivier Morales, Samir Acherar, Celine Frochot, Nadira Delhem

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Ovarian cancer (OC) is one of the most defying diseases in gynecologic oncology. Even though surgery remains crucial in the therapy of patients with primary ovarian cancer, recurrent recidivism calls for the development of new therapy protocols to propose for patients dealing with this cancer. FRα is described as a tumor‐associated antigen in OC, where FRα expression is usually linked with more poorly differentiated, aggressive tumors. The Photodynamic treatment (PDT) available data have shown improvements in the uptake of small tumors and in the induction of a proper anti-tumoral immune response. In order to target specifically peritoneal metastatis, which overexpress FRα, a new-patented PS coupled with folic acid has been developed in our team. Herein we propose PDT using this new patented PS for PDT applied in an in vivo mice model. The efficacy of the treatment was evaluated in mice without and with PBMC reconstitution. Mice were divided into four groups: Non-Treated, PS, Light Only, and PDT Treated and subjected to illumination by laser set at 668nm with a duration of illumination of 45 minutes (or 1 min of illumination followed by 2 minutes of pause repeated 45 times). When mice were not reconstituted and after fractionized PDT protocol, a significant decrease in the tumor volume was noticed. An induction in the anti-tumoral cytokine IFNγ chaperoned this decrease while a subsequent inhibition in the cytokine TGFβ. Even more crucial, when mice were reconstituted and upon PDT, the fold of tumor decrease was even higher. An immune response was activated decoded with an increase in NK, CD3 +, LT helper and Cytotoxic T cells. Thereafter, an increase in the expression of the cytokines IFNγ and TNFα were noticed while an inhibition in TGFβ, IL8 and IL10 accompanied this immune response activation. Therefore, our work has shown for the first time that a fractionized PDT protocol using a folate-targeted PDT is effective for treatment of ovarian cancer. The interest in using PDT in this case, goes beyond the local induction of tumor apoptosis only, but can promote subsequent anti-tumor response. Most of the therapies currently used to treat ovarian cancer, have an uncooperative outcomes on the host immune response. The readiness of a tumor adjuvant treatment like PDT adequate in eliminating the tumor and in concert stimulating anti-tumor immunity would be weighty.

Keywords: folate receptor, ovarian cancer, photodynamic therapy, humanized mice model

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Authors: Diom Loreen Ndum, Omarine Njimanted

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With increasing automation in clinical laboratories, the requirements for quality control materials have greatly increased in order to monitor daily performance. The constant use of commercial control material is not economically feasible for many developing countries because of non-availability or the high-cost of the materials. Therefore, preparation and use of in-house quality control serum will be a very cost-effective measure with respect to laboratory needs.The objective of this study was to determine the efficacy of in-house prepared pooled sera with respect to commercially acquired control sample for routine internal quality control at the Nkwen District Hospital Laboratory. This was an analytical study, serum was taken from leftover serum samples of 5 healthy adult blood donors at the blood bank of Nkwen District Hospital, which had been screened negative for human immunodeficiency virus (HIV), hepatitis C virus (HCV) and Hepatitis B antigens (HBsAg), and were pooled together in a sterile container. From the pooled sera, sixty aliquots of 150µL each were prepared. Forty aliquots of 150µL each of commercially acquired samples were prepared after reconstitution and stored in a deep freezer at − 20°C until it was required for analysis. This study started from the 9th June to 12th August 2022. Every day, alongside with commercial control sample, one aliquot of pooled sera was removed from the deep freezer and allowed to thaw before analyzed for the following parameters: blood urea, serum creatinine, aspartate aminotransferase (AST), alanine aminotransferase (ALT), potassium and sodium. After getting the first 20 values for each parameter of pooled sera, the mean, standard deviation and coefficient of variation were calculated, and a Levey-Jennings (L-J) chart established. The mean and standard deviation for commercially acquired control sample was provided by the manufacturer. The following results were observed; pooled sera had lesser standard deviation for creatinine, urea and AST than commercially acquired control samples. There was statistically significant difference (p<0.05) between the mean values of creatinine, urea and AST for in-house quality control when compared with commercial control. The coefficient of variation for the parameters for both commercial control and in-house control samples were less than 30%, which is an acceptable difference. The L-J charts revealed shifts and trends (warning signs), so troubleshooting and corrective measures were taken. In conclusion, in-house quality control sample prepared from pooled serum can be a good control sample for routine internal quality control.

Keywords: internal quality control, levey-jennings chart, pooled sera, shifts, trends, westgard rules

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Authors: Aberdam Edith, Sangari Linda, Petit Isabelle, Aberdam Daniel

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Background and Rationale: Transparent avascularised cornea is essential for normal vision and depends on limbal stem cells (LSC) that reside between the cornea and the conjunctiva. Ocular burns or injuries may destroy the limbus, causing limbal stem cell deficiency (LSCD). The cornea becomes vascularised by invaded conjunctival cells, the stroma is scarring, resulting in corneal opacity and loss of vision. Grafted autologous limbus or cultivated autologous LCS can restore the vision, unless the two eyes are affected. Alternative cellular sources have been tested in the last decades, including oral mucosa or hair follicle epithelial cells. However, only partial success has been achieved by the use of these cells since they were not able to uniformly commit into corneal epithelial cells. Human pluripotent stem cells (iPSC) display both unlimited growth capacity and ability to differentiate into any cell type. Our goal was to design a standardized and reproducible protocol to produce transplantable autologous LSC from patients through cell reprogramming technology. Methodology: First, keratinocyte primary culture was established from a small number of plucked hair follicles of healthy donors. The resulting epithelial cells were reprogrammed into induced pluripotent stem cells (iPSCs) and further differentiate into corneal epithelial cells (CEC), according to a robust protocol that recapitulates the main step of corneal embryonic development. qRT-PCR analysis and immunofluorescent staining during the course of differentiation confirm the expression of stage specific markers of corneal embryonic lineage. First appear ectodermal progenitor-specific cytokeratins K8/K18, followed at day 7 by limbal-specific PAX6, TP63 and cytokeratins K5/K14. At day 15, K3/K12+-corneal cells are present. To amplify the iPSC-derived LSC (named COiPSC), intact small epithelial colonies were detached and cultivated in limbal cell-specific medium. In that culture conditions, the COiPSC can be frozen and thaw at any passage, while retaining their corneal characteristics for at least eight passages. To evaluate the potential of COiPSC as an alternative ocular toxicity model, COiPSC were treated at passage P0 to P4 with increasing amounts of SDS and Benzalkonium. Cell proliferation and apoptosis of treated cells was compared to LSC and the SV40-immortalized human corneal epithelial cell line (HCE) routinely used by cosmetological industrials. Of note, HCE are more resistant to toxicity than LSC. At P0, COiPSC were systematically more resistant to chemical toxicity than LSC and even to HCE. Remarkably, this behavior changed with passage since COiPSC at P2 became identical to LSC and thus closer to physiology than HCE. Comparative transcriptome analysis confirmed that COiPSC from P2 are similar to a mixture of LSC and CEC. Finally, by organotypic reconstitution assay, we demonstrated the ability of COiPSC to produce a 3D corneal epithelium on a stromal equivalent made of keratocytes. Conclusion: COiPSC could become valuable for two main applications: (1) an alternative robust tool to perform, in a reproducible and physiological manner, toxicity assays for cosmetic products and pharmacological tests of drugs. (2). COiPSC could become an alternative autologous source for cornea transplantation for LSCD.

Keywords: Limbal stem cell deficiency, iPSC, cornea, limbal stem cells

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