Gonadihypoplasia lapinlehmillä Ruotsissa ja Suomessa

Tämän lisensiaatintutkielman tavoitteena on selvittää pohjoissuomenkarjan naudoilla esiintyvän gonadien hypoplasian eli sukurauhasten vajaakehittyneisyyden syitä. Sukurauhasten vajaakehittyneisyyden tiedetään johtuvan itusolujen puutteesta sukurauhasissa. Itusolujen puutos on havaittavissa jo varhaisessa sikiönkehityksen vaiheessa ja sen epäillään johtuvan häiriöstä alkuitusolujen soluvaelluksessa. KIT-reseptorin (tunnetaan myös nimellä Ckit tai C-Kit) tiedetään olevan merkittävässä roolissa alkuitusolujen soluvaelluksessa ja nykyisten geenitutkimusmenetelmien avulla pyrittiin selvittämään sen osuutta pohjoissuomenkarjan sukurauhasten vajaakehittyneisyydessä. Yhteensä 303 pohjoissuomenkarjan nautaa tutkittiin kliinisesti sukurauhasten vajaakehittyneisyyden varalta. Sonnien ja sonnivasikoiden tutkimus suoritettiin mittaamalla ja tunnustelemalla niiden kivekset. Lehmillä ja hiehoilla munasarjat tutkittiin rektaalisesti palpoimalla. Lisäksi osalta eläimiä otettiin 9 ml laskimoverta EDTA-putkeen DNA-tutkimusta varten. Lehmillä ja hiehoilla todettiin toinen tai molemmat munasarjat vajaakehittyneiksi, mikäli niiden koko oli hyvin pieni tai ne olivat mahdottomat löytää rektaalitutkimuksessa. Sonneilla kives todettiin vajaakehittyneeksi, jos sen koko oli korkeintaan kolmasosa normaalin kiveksen koosta. Sonnivasikoilla kives todettiin vajaakehittyneeksi, jos sen koko oli korkeintaan puolet normaalin kiveksen koosta. Lisäksi 42 eläimen sukurauhaset tutkittiin teurastuksen tai kliinisen kastraation jälkeen. Vajaakehittyneeksi sukurauhaset todettiin aiemmin kuvatun mukaisesti ja histologisten leikkeiden avulla, jos niistä ei löytynyt sukusoluja tai niiden esiasteita. DNA-näytteet genotyypitettiin 96 naudan osalta Illumina BovineHD sirun ja Illumina's Beadstudio ohjelmiston avulla. Koko genominlaajuista assosiaatiotutkimusta (GWAS) varten naudat jaettiin ryhmiin sairauden ja värityksen mukaan. Tutkituista 345 pohjoissuomenkarjan naudoista 16 eläimellä oli hypoplastiset sukurauhaset. Genotyyppaustulosten mukaan kromosomi 29 oli vahvasti yhteydessä sukurauhasten vajaakehittyneisyyteen ja kopioluvun vaihteluita (CNV) löytyi kaikkiaan 2101 kappaletta ja kromosomista 6 löydetty CNV sisälsi KIT-geenin kokonaisuudessaan. Toinen kromosomista 29 löydetty CNV-segmentti oli GWAS-tutkimuksessa löydetyn alueen välittömässä läheisyydessä. Kromosomista 29 löydetty ektooppinen KIT-geeni liittyy sukurauhasten vajaakehitykseen pohjoissuomenkarjan naudoilla. Löydös on hypoteesin mukainen, mutta tarkkaa sukurauhasten vajaakehityksen syytä ei tutkimuksen tuloksista pystytä määrittelemään

An miR-200 Cluster on Chromosome 23 Regulates Sperm Motility in Zebrafish

Besides its well-documented roles in cell proliferation, apoptosis, and carcinogenesis, the function of the p53-microRNA axis has been recently revealed in the reproductive system. Recent studies indicated that miR-200 family members are dysregulated in nonobstructive azoospermia patients, whereas their functions remain poorly documented. The aim of this study was to investigate the function of the miR-200 family on zebrafish testis development and sperm activity. There was no substantial difference in testis morphology and histology between wild-type (WT) and knockout zebrafish with deletion of miR-200 cluster on chromosome 6 (chr6-miR-200-KO) or on chromosome 23 (chr23-miR-200-KO). Interestingly, compared with WT zebrafish, the chr6-miR-200-KO zebrafish had no difference on sperm motility, whereas chr23-miR-200-KO zebrafish showed significantly improved sperm motility. Consistently, ectopic expression of miR-429a, miR-200a, and miR-200b, which are located in the miR-200 cluster on chromosome 23, significantly reduced motility traits of sperm. Several sperm motility-related genes, such as amh, wt1a, and srd5a2b have been confirmed as direct targets of miR-200s on chr23. 17a-ethynylestradiol (EE2) exposure resulted in upregulated expression of p53 and miR-429a in testis and impairment of sperm motility. Strikingly, in p53 mutant zebrafish testis, the expression levels of miR-200s on chr23 were significantly reduced and accompanied by a stimulation of sperm motility. Moreover, the upregulation of miR-429a associated with EE2 treatment was abolished in testis with p53 mutation. And the impairment of sperm activity by EE2 treatment was also eliminated when p53 was mutated. Together, our results reveal that miR-200 cluster on chromosome 23 controls sperm motility in a p53-dependent manner.</p

The Role of c-Kit Receptor Tyrosine Kinase in Beta-Cell Proliferation, Function and Survival

c-Kit, a receptor tyrosine kinase, interacts with Stem Cell Factor (SCF), mediating cell differentiation, function, and survival. c-Kit is critical for the development and maintenance of beta-cell function in both rodents and humans. The mutation of c-Kit at W locus (c-KitWv/+) in mice results in an early onset of diabetes. However, the underlying mechanisms by which c-Kit deficiency leads to beta-cell failure are unknown. Therefore, studying SCF/c-Kit downstream signaling pathways is essential to understanding the precise mechanism by which c-Kit regulates beta-cell survival and function in vivo. We identified that dysregulated Akt/Glycogen synthase kinase 3β (Gsk3β)/cyclin D1 pathway, downstream of c-Kit, is responsible for reduced beta-cell proliferation, leading to a severe loss of beta-cell mass in c-KitWv/+ mice. An up-regulation of Fas-mediated caspase-dependent apoptotic machinery is also associated with beta-cell death in c-KitWv/+ mouse islets. The loss of functional Fas (lpr mutation) reversed beta-cell apoptosis and dysfunction in c-KitWv/+;Faslpr/lpr double mutant mice, demonstrating that a balance between c-Kit and Fas signaling is critical for beta-cell survival and function. To further delineate the primary functional role of c-Kit in beta-cells, we developed a transgenic (c-KitβTg) mouse model with beta-cell specific c-KIT overexpression. c-KitβTg mice exhibited increased beta-cell mass with improved insulin secretion, which is mediated by up-regulation of Akt/Gsk3β/cyclin D1 pathway. c-KIT overexpression in beta-cells not only protected islet function from 4 weeks of high-fat-diet (HFD) challenge, but also recused the onset of diabetes observed in c-KitWv/+ mice. We also found that c-Kit signaling plays a critical role in islet vascularization. c-Kit mediates VEGF-A production via the Akt/mTOR pathway in vivo. c-KIT overexpression in beta-cells rescued the islet vascular defects in c-KitWv/+ mice. However, under long-term HFD challenge, c-KitβTg mouse islets displayed dilated vessels with reduced beta-cell mass and increased beta-cell apoptosis. The observed beta-cell failure was likely associate with expanded islet vasculature causing increased islet inflammatory response. In conclusion, this series of studies represent an integrated in vitro and in vivo approach aimed at unraveling the cellular mechanisms by which SCF/c-Kit regulates beta-cell survival and function

Germ Cell Tumor

The book aims to provide an overview of current knowledge regarding germ cell tumors. It deals with the clinical presentations, treatment modalities, the biology and genetics of germ cell tumors in children and adults. Most chapters are focused on testicular germ cell tumors whose incidence has been increasing in young males. Included are reviews on the pathogenesis, risk factors, diagnosis and treatment regimens applied to precursor, pre-invasive lesions as well as to seminomatous and non-seminomatous germ cell tumors of the testes. In addition, a review is included on the diagnosis and current management options for intracranial germ cell tumors in children. Authors have also contributed articles on the genetics and epigenetics of germ cell tumor development in humans and in the mouse model system. This book will be of interest to scientists, physicians and lay readers wishing to review recent developments in the field of germ cell cancers

Antioxidant and DPPH-Scavenging Activities of Compounds and Ethanolic Extract of the Leaf and Twigs of Caesalpinia bonduc L. Roxb.

Antioxidant effects of ethanolic extract of Caesalpinia bonduc and its isolated bioactive compounds were evaluated in vitro. The compounds included two new cassanediterpenes, 1α,7α-diacetoxy-5α,6β-dihydroxyl-cass-14(15)-epoxy-16,12-olide (1)and 12α-ethoxyl-1α,14β-diacetoxy-2α,5α-dihydroxyl cass-13(15)-en-16,12-olide(2); and others, bonducellin (3), 7,4’-dihydroxy-3,11-dehydrohomoisoflavanone (4), daucosterol (5), luteolin (6), quercetin-3-methyl ether (7) and kaempferol-3-O-α-L-rhamnopyranosyl-(1Ç2)-β-D-xylopyranoside (8). The antioxidant properties of the extract and compounds were assessed by the measurement of the total phenolic content, ascorbic acid content, total antioxidant capacity and 1-1-diphenyl-2-picryl hydrazyl (DPPH) and hydrogen peroxide radicals scavenging activities.Compounds 3, 6, 7 and ethanolic extract had DPPH scavenging activities with IC50 values of 186, 75, 17 and 102 μg/ml respectively when compared to vitamin C with 15 μg/ml. On the other hand, no significant results were obtained for hydrogen peroxide radical. In addition, compound 7 has the highest phenolic content of 0.81±0.01 mg/ml of gallic acid equivalent while compound 8 showed the highest total antioxidant capacity with 254.31±3.54 and 199.82±2.78 μg/ml gallic and ascorbic acid equivalent respectively. Compound 4 and ethanolic extract showed a high ascorbic acid content of 2.26±0.01 and 6.78±0.03 mg/ml respectively.The results obtained showed the antioxidant activity of the ethanolic extract of C. bonduc and deduced that this activity was mediated by its isolated bioactive compounds