MicroRNAs in the bovine ovary and placentas derived from in vivo, in vitro and nuclear transfer pregnancies

MicroRNAs are the major class of gene regulating molecules playing pivotal roles at post-transcriptional level. Identification and expression profiling are the initial steps to understand their regulation of biological processes. Despite increasing efforts in miRNAs characterization in different species, little is known in the bovine reproductive tissues especially in ovary and placenta. Two subsequent studies were carried out to the expression of miRNAs in bovine ovary and Day-50 placenta derived from different sources of pregnancy. The first study aims to identify and characterize miRNAs in bovine ovary through cloning, expression analysis and target prediction. The constructed miRNA library revealed cloning of 50 known and 24 novel miRNAs. Among these, 38 were new miRNAs which were derived from 43 distinct loci with characteristic secondary structure. Most of the miRNAs were cloned multiple times and thereby reflecting their expression level and potential role in the ovary. Analysis of identified miRNAs in different intra-ovarian structures and other tissues reveals their stage and tissue specific expression patterns. Furthermore, in silico target prediction and Gene Ontology analysis of the targets genes identified several biological processes and pathways underlying the ovarian function. Results of this study suggest the presence of miRNAs in the bovine ovary; thereby elucidate their potential role in regulating diverse mechanisms underlying the ovarian functionality. The second study aimed to elucidate the difference in expression profile of miRNAs in the placenta at day 50 derived from Somatic cell nuclear transfer (SCNT), in vitro production (IVP) and artificial insemination (AI) pregnancies by quantifying 377 miRNAs. The study reveals a massive deregulation of miRNAs which were poorly reprogrammed and affected as large chromosomal cluster as well as miRNA families in the NT and IVP placenta compared to that of AI. Furthermore, cell specific localization miRNAs in the expanded blastocysts and expression profiling in different developmental stages of embryos and placenta identified that the major deregulation of miRNAs arises at day 50 of NT and IVP pregnancies. This deregulation were found to be less dependent on global DNA methylation, rather aberrant miRNA processing molecules were evidenced. Among them, observed down regulation of AGO2 could be a reason for global down regulation of miRNAs in the NT or IVP placenta. Identified deregulation of miRNAs might associate to the abnormal placentogenesis in NT or IVP pregnancies, which are the results of aberrant genetic and epigenetic modification. Result of this study will help to move one step closer towards improving the efficiency of nuclear transfer pregnancy. Altogether, the present study has discovered miRNAs in the bovine ovary and elucidated the pattern of expression of miRNAs along with their regulatory mechanism in the placenta derived from pregnancies of various origins.Untersuchung von MicroRNAs in bovinen Ovarien und Plazenten aus geklonten, in vivo und in vitro erzeugten Trächtigkeiten MicroRNAs gehören zur großen Klasse der genregulierenden Moleküle und spielen eine bedeutende Rolle auf der posttranskriptionellen Ebene. Die Identifikation und die Erstellung von Expressionsprofilen sind die ersten Schritte für ein besseres Verständnis der miRNA und ihrer Beteiligung an der Regulierung von biologischen Prozessen. Trotz der zunehmenden Charakterisierung der miRNA in verschiedenen Spezies, sind kaum Untersuchungen in bovinen Ovarien und Plazenten bekannt. In zwei Studien wurde die miRNA Expression in bovinen Ovarien und in Plazenten am Tag 50 der Trächtigkeit, in unterschiedlich erzeugten Schwangerschaften untersucht. Das Ziel der ersten Studie war die Identifizierung und Charakterisierung von miRNAs in klonierten bovinen Ovarien, Expressionsanalysen sowie Target-Vorhersagen. Die konstruierte miRNA-Bibliothek ergab 50 bekannte und 24 neue miRNAs. Unter diesen waren 38 neue bovine miRNAs die von 43 eindeutigen Loci abgeleitet werden konnten, die eine charakteristische sekundäre Struktur zeigten. Durch die mehrfache Klonierung der meisten miRNAs, konnte ihr Expressionsniveau in den Ovarien erfasst werden. Analysen von miRNAs in unterschiedlichen intra-ovariellen Strukturen sowie anderen Geweben zeigten ihr phasen- und gewebsspezifisches Expressionsmuster. Des Weiteren konnte durch bioinformatische Auswertungen und Gen Ontology Analysen der Target Gene verschiedene biologische Prozesse und Pathways der Ovarienfunktion identifiziert werden. Die Ergebnisse dieser Studie deuten auf das Vorhandensein von miRNAs in bovinen Ovarien hin und verdeutlichen die potenzielle Bedeutung in der Regulierung diverser Mechanismen der Ovarienfunktion. Die zweite Studie sollte die Unterschiede von Expressionsprofilen von miRNAs in Plazenten am Tag 50 der Trächtigkeit von SCNT, IVP und AI Schwangerschaften durch Quantifizierung von 377 miRNAs aufklären. Die Studie zeigte eine starke reduzierte Expression und eine geringe Reprogrammierung der miRNAs in NT und IVP im Vergleich zu AI. Diese miRNAs gehören vermutlich zu einer miRNA Familie in einem chromosomalen Cluster. Des Weiteren zeigten zellspezifische Lokalisationen von miRNAs in der expandierenden Blastozyste und Expressionsprofile von unterschiedlichen Entwicklungsstadien im Embryo und in der Plazenta, dass die wichtigsten Deregulierungen von miRNAs am Tag 50 der Trächtigkeit in NT und IVP entstehen. Diese Deregulierung erwies sich als weniger abhängig von einer globalen DNA-Methylierung, vielmehr wurde eine Abweichung in den miRNA-Prozessgenen belegt. Von diesen könnte die reduzierte Expression von AGO2 die Ursache für die globale Deregulierung der miRNAs in NT oder IVP Plazenten sein. Die identifizierten Deregulationen der miRNAs könnten im Zusammenhang mit abnormaler Plazentogenese in NT oder IVP Trächtigkeiten stehen. Die Ursachen für Abnormalitäten in der Plazentogenese liegen in genetischen und epigenetischen Modifikationen. Zusammenfassend konnte durch diese Studien miRNAs in bovinen Ovarien identifiziert werden und Expressionsmustern der miRNAs sowie ihre regulierenden Mechanismen in der Plazenta von unterschiedlichen Trächtigkeiten beschrieben werden

Informed Consent and the Patients of Bangladesh

In medical practice, informed consent plays a vital role in making ethical decisions. In recent years, informed consent also appears to be a key issue in biomedical discussion. Several definitions of informed consent have already been proposed. However, the socio-economic conditions of different countries are not the same. Many countries differ culturally as well as in literacy rate. Therefore, a unified concept of informed consent might not be justified in every context. This paper discusses the importance of informed consent, different views regarding informed consent and the limitations of applying one single idea of informed consent in every context. It argues that the most plausible concept of informed consent which is achievable for developing countries, like Bangladesh, is based on care ethics. The paper concludes that informed consent could be a ‘natural outcome’ and is really not a barrier in the patient-physician relationship if the idea is seen from the perspective of care ethics

Identification, Self-Realization and Spirituality: A Comparative Study of Environmental Philosophy

Ph.DDOCTOR OF PHILOSOPH

Exploring lncRNAs associated with human pancreatic islet cell death induced by transfer of adoptive lymphocytes in a humanized mouse model

BackgroundLong noncoding RNA (lncRNA)-mediated posttranscriptional and epigenetic landscapes of gene regulation are associated with numerous human diseases. However, the regulatory mechanisms governing human β-cell function and survival remain unknown. Owing to technical and ethical constraints, studying the direct role of lncRNAs in β-cell function and survival in humans in vivo is difficult. Therefore, we utilized humanized mice with human islets to investigate lncRNA expression using whole transcriptome shotgun sequencing. Our study aimed to characterize lncRNAs that may be crucial for human islet cell function and survival.MethodsHuman β-cell death was induced in humanized mice engrafted with functional human islets. Using these humanized mice harboring human islets with induced β-cell death, we investigated lncRNA expression through whole transcriptome shotgun sequencing. Additionally, we systematically identified, characterized, and explored the regulatory functions of lncRNAs that are potentially important for human pancreatic islet cell function and survival.ResultsHuman islet cell death was induced in humanized mice engrafted with functional human islets. RNA sequencing analysis of isolated human islets, islet grafts from humanized mice with and without induced cell death, revealed aberrant expression of a distinct set of lncRNAs that are associated with the deregulated mRNAs important for cellular processes and molecular pathways related to β-cell function and survival. A total of 10 lncRNA isoforms (SCYL1-1:22, POLG2-1:1, CTRB1-1:1, SRPK1-1:1, GTF3C5-1:1, PPY-1:1, CTRB1-1:5, CPA5-1:1, BCAR1-2:1, and CTRB1-1:4) were identified as highly enriched and specific to human islets. These lncRNAs were deregulated in human islets from donors with different BMIs and with type 2 diabetes (T2D), as well as in cultured human islets with glucose stimulation and induced cell death induced by cytokines. Aberrant expression of these lncRNAs was detected in the exosomes from the medium used to culture islets with cytokines.ConclusionIslet-enriched and specific human lncRNAs are deregulated in human islet grafts and cultured human islets with induced cell death. These lncRNAs may be crucial for human β-cell function and survival and could have an impact on identifying biomarkers for β-cell loss and discovering novel therapeutic targets to enhance β-cell function and survival

Critical factors for optimum biodegradation of bast fiber’s gums in bacterial retting

Bast fiber plants require a post-harvest process to yield useable natural cellulosic fibers, denoted as retting or degumming. It encompasses the degradation of the cell wall’s non-cellulosic gummy substances (NCGs), facilitating fibers separations, setting the fiber’s quality, and determining downstream usages. Due to the inconvenience of traditional retting practices, bacterial inoculum and enzyme applications for retting gained attention. Therefore, concurrent changes of agroclimatic and socioeconomic conditions, the conventional water retting confront multiple difficulties, bast industries become vulnerable, and bacterial agents mediated augmented bio-retting processes trying to adapt to sustainability. However, this process’s success demands a delicate balance among substrates and retting-related biotic and abiotic factors. These critical factors were coupled to degrade bast fibers NCGs in bacterial retting while holistically disregarded in basic research. In this study, a set of factors were defined that critically regulates the process and requires to be comprehended to achieve optimum retting without failure. This review presents the bacterial strain characteristics, enzyme potentials, specific bast plant cell wall’s structure, compositions, solvents, and interactions relating to the maximum NCGs removal. Among plants, associated factors pectin is the primary biding material that determines the process’s dynamics, while its degree of esterification has a proficient effect through bacterial enzymatic degradation. The accomplished bast plant cell wall’s structure, macerating solvents pH, and temperature greatly influence the bacterial retting process. This article also highlights the remediation process of water retting pollution in a biocompatible manner concerning the bast fiber industry’s endurance

Isolation of Alkalophilic Pectinolytic Bacteria and their Bio Retting Effect on Kenaf Fiber Compositions

Retting is the most limiting process of high-quality cellulosic kenaf bast fiber production which facilitating the separation of useable fiber from the plants' cell wall matrix. Existing traditional water retting approach confronts ineptitude and eutrophication related complications. Aiming to enhance the kenaf bio-retting process, sixty-seven alkalophilic bacterial colonies were isolated from paddy land soil sediments and kenaf retting water. These isolates were subsequently screened, of that two isolates were selected based on hyper qualitative and quantitative pectinolytic enzymatic measures. 16s rDNA gene sequence analysis revealed that both two strains were closely related to Bacillus pumilus species and designated as KRB56 and KRB22. These strains were applied in augmented non-sterile kenaf tank retting to investigate their kenaf retting efficiency and yielded fiber were analyzed for chemical compositions. Results revealed that, stains KRB56 and KRB22 significantly improve the retting process by degradation of 82.78% and 75.28% non-cellulosic gums, respectively comparing with uninoculated treatment niche (62.12%). These bacterial treated fiber samples showed thinner, smooth, and cleaner fibers surface morphology by SEM indicates sufficient non-cellulosic gums (NCGs) removal comparing with URKF. Moreover, yielded fibers were examined for chemical composition, FTIR, XRD test. Results revealed that compare to un retted and un inoculated kenaf fiber, bacterial treated kenaf fiber increases cellulose portions, and their crystallinity index increases 35.50-41.30 % due to sufficient NCGs removal. This study's findings indicate that isolated alkalophilic bacterial strains KRB56 and KRB22 were effectively to be used as kenaf bio retting agents to produce quality kenaf fiber

Assessment of eye health care services of Bangladesh using eye care service assessment tools

Background: Bangladesh is being the commissioner for oaths to vision 2020, a global campaign for elimination of avoidable blindness by 2020, formulated a national eye care plan. This report illustrates the present status of Bangladesh eye health care service using eye care service assessment tool (ECSAT) that assesses an eye health system across six ‘building blocks’ of a health system.Methods: The study followed a mixed method to collect data. World health organization (WHO) standard ECSAT was used to gather information on eye care service. A purposive sampling method was used. Data from the assessment were extracted and all the information was cross-checked with leading stakeholders of ministry of health.Results: Eye care planning is led by the national eye care. There is a national eye health action plan and a national eye health coordination office under the ministry of health. The health delivery system includes primarily government and non-profit facilities with eight hospitals delivering specialist eye care services across the country. A significant proportion of eye care is provided through community outreach camps and a network of primary and community health workers. The national cataract surgical rate (CSR) is estimated at 2600 per million populations per year.Conclusions: This assessment suggests that although Bangladesh has made some progress towards elimination of avoidable blindness, it would be difficult to retain without further significant investment with a transparent accountability framework in eye health considering all limitation and contemporary challenges

Augmented Retting Effect on Kenaf Fibers Using Alkalophilic Pectinase-Producing Bacteria in Combination with Water Solvents

A degumming approach is used in this paper with alkalophilic pectinase-producing bacteria (APPB) and two sources of water solvents to address the existing conventional water retting complexities of kenaf. The incorporation of APPB was confirmed based on their retting feasibilities and multiple cell-wall-degrading enzymatic delicacy. The combinations of APPB with seawater offered retting achievements within six-day retting in non-sterile conditions. These retting niches showed maximum (14.67 U/mL) pectinase activity with fiber separation feasibilities of 4.75 Fried test score. The yielded fiber composition analysis showed a higher cellulose composition (84.65%) and the least amount of hemicellulose, pectin, and ligneous gummy substances. The transmission electron microscopy scan of the yielded fibers showed smooth fiber surfaces, 84.20 µm fiber diameter, and 7.65 g/tex fine fiber compared with uninoculated and combinations of freshwater treatments. The FTIR spectra revealed the cellulosic discrepancies of the retting treatments by monitoring O-H and C=O stretching at ~3300 cm−1 and ~1730 cm−1 wavenumbers. These findings are compelling to yield kenaf fibers of quality considering the existing retting difficulties

Identification and characterization of miRNAs expressed in the bovine ovary

<p>Abstract</p> <p>Background</p> <p>MicroRNAs are the major class of gene-regulating molecules playing diverse roles through sequence complementarity to target mRNAs at post-transcriptional level. Tightly regulated expression and interaction of a multitude of genes for ovarian folliculogenesis could be regulated by these miRNAs. Identification of them is the first step towards understanding miRNA-guided gene regulation in different biological functions. Despite increasing efforts in miRNAs identification across various species and diverse tissue types, little is known about bovine ovarian miRNAs. Here, we report the identification and characterization of miRNAs expressed in the bovine ovary through cloning, expression analysis and target prediction.</p> <p>Results</p> <p>The miRNA library (5'-independent ligation cloning method), which was constructed from bovine ovary in this study, revealed cloning of 50 known and 24 novel miRNAs. Among all identified miRNAs, 38 were found to be new for bovine and were derived from 43 distinct loci showing characteristic secondary structure. While 22 miRNAs precursor loci were found to be well conserved in more than one species, 16 were found to be bovine specific. Most of the miRNAs were cloned multiple times, in which let-7a, let-7b, let-7c, miR-21, miR-23b, miR-24, miR-27a, miR-126 and miR-143 were cloned 10, 28, 13, 4, 11, 7, 6, 4 and 11 times, respectively. Expression analysis of all new and some annotated miRNAs in different intra-ovarian structures and in other multiple tissues showed that some were present ubiquitously while others were differentially expressed among different tissue types. Bta-miR-29a was localized in the follicular cells at different developmental stages in the cyclic ovary. Bio-informatics prediction, screening and Gene Ontology analysis of miRNAs targets identified several biological processes and pathways underlying the ovarian function.</p> <p>Conclusion</p> <p>Results of this study suggest the presence of miRNAs in the bovine ovary, thereby elucidate their potential role in regulating diverse molecular and physiological pathways underlying the ovarian functionality. This information will give insights into bovine ovarian miRNAs, which can be further characterized for their role in follicular development and female fertility as well.</p