Vitamin D receptor-targeted treatment to prevent pathological dedifferentiation of pancreatic ß cells under hyperglycaemic stress.

Dedifferentiation has been identified as one of the causes of ß-cell failure resulting in type 2 diabetes (T2D). This study tested whether increasing vitamin D receptor (VDR) expression prevents dedifferentiation of ß cells in a high-glucose state in vitro. Culturing a mouse insulinoma cell line (MIN6) in a high-glucose environment decreased VDR expression. However, increased VDR following vitamin D3 (VD3) treatment improved insulin release of early-passage MIN6 and insulin index of db/- (heterozygous) islets to levels seen in normal functional islets. Treatment with VD3, its analogues and derivatives also increased the expression of essential transcription factors, such as Pdx1, MafA and VDR itself, ultimately increasing expression of Ins1 and Ins2, which might protect ß cells against dedifferentiation. VD3 agonist lithocholic acid (LCA) propionate was the most potent candidate molecule for protecting against dedifferentiation, and an e-pharmacophore mapping model confirmed that LCA propionate exhibits a stabilizing conformation within the VDR binding site. This study concluded that treating db/+ islets with a VD3 analogue and/or derivatives can increase VDR activity, preventing the pathological dedifferentiation of ß cells and the onset of T2D.

Incomplete Re-Expression of Neuroendocrine Progenitor/Stem Cell Markers is a Key Feature of ß-Cell Dedifferentiation.

There is increasing evidence to suggest that type 2 diabetes mellitus (T2D), a pandemic metabolic disease, may be caused by ß-cell dedifferentiation (ßCD). However, there is currently no universal definition of ßCD, and the underlying mechanism is poorly understood. We hypothesise that a high-glucose in vitro environment mimics hyperglycaemia in vivo and that ß cells grown in this milieu over a long period will undergo dedifferentiation. In the present study, we report that the pancreatic ß cell line mouse insulinoma 6 (MIN6) grown under a high-glucose condition did not undergo massive cell death but exhibited a glucose-stimulated insulin-secreting profile similar to that of immature ß cells. The expression of insulin and the glucose-sensing molecule glucose transporter 2 (Glut2) in late passage MIN6 cells was significantly lower than the early passage at both the RNA and protein levels. Mechanistically, these cells also expressed significantly less of the 'pancreatic and duodenal homebox1' (Pdx1) ß-cell transcription factor. Finally, passaged MIN6 cells dedifferentiated to demonstrate some features of ß-cell precursors, as well as neuroendocrine markers, in addition to expressing both glucagon and insulin. Thus, we concluded that high-glucose passaged MIN6 cells passaged MIN6 cells. provide a cellular model of ß-cell dedifferentiation that can help researchers develop a better understanding of this process. These findings provide new insights that may enhance knowledge of the pathophysiology of T2D and facilitate the establishment of a novel strategy by which this disease can be treated.

Omentin-1 plasma levels and omentin-1 expression are decreased in psoriatic lesions of psoriasis patients.

Previous studies have suggested that psoriasis is associated with individuals who are overweight or obese. Omentin is a recently discovered adipokine that is involved in chronic inflammatory processes. This study evaluated the relationship between omentin and psoriasis, focusing on omentin-1 serum levels, skin expression of omentin-1, and omentin-1 gene polymorphism (rs2274907; Val109Asp). Levels of omentin-1 in serum samples from 44 patients with psoriasis and 38 healthy controls were analyzed by enzyme-linked immunosorbent assay. The expressions of omentin-1 were measured by immunohistochemistry in 3 psoriasis affected skins and 3 normal skins. The rs2274907 variant was typed using a SNaPshot assay in 354 cases and 214 controls. We found significantly lower serum levels of omentin-1 in psoriasis patients compared with healthy controls (p < 0.001) with an inverse correlation with the Psoriasis Area and Severity Index (p < 0.01). The comparison of genotype and allele distribution revealed no significant difference in genotype frequency between psoriasis patients and controls. Therefore, omentin-1 may have a role in the pathogenesis of psoriasis and might be a potential biological marker for psoriasis severity.

A mutation in SART3 gene in a Chinese pedigree with disseminated superficial actinic porokeratosis.

BACKGROUND: Disseminated superficial actinic porokeratosis (DSAP) is an uncommon autosomal dominant chronic disorder of keratinization, characterized by multiple superficial keratotic lesions surrounded by a slightly raised keratotic border. Thus far, although two loci for DSAP have been identified, and the genetic basis and pathogenesis of this disorder have not been elucidated. OBJECTIVES: To determine the locus of DSAP and identify the candidate gene(s) of the disease. METHODS: Genome-wide scan and linkage analysis were performed in a six-generation Chinese family with DSAP. The coding exons of the candidate genes were sequenced to analyse and detect the nucleotide variations. RESULTS: Linkage analysis showed that the maximum two-point lod score of 5.56 was obtained with the marker D12S79 at a recombination fraction theta of 0.00. Haplotype analysis defined the critical region for DSAP between D12S330 and D12S1612 on 12q24.1-24.2. By sequence analysis, we found a Val591Met mutation in SART3 in all affected individuals of the family. CONCLUSION: SART3 is a candidate gene for DSAP, and is possibly involved in the pathogenesis of DSAP.

Regulation of laminin 1-induced pancreatic beta-cell differentiation by alpha6 integrin and alpha-dystroglycan.

BACKGROUND: The ability to manipulate the development of pancreatic insulin-producing beta cells has implications for the treatment of type 1 diabetes. Previously, we found that laminin-1, a basement membrane trimeric glycoprotein, promotes beta-cell differentiation. We have investigated the mechanism of this effect, using agents that block the receptors for laminin-1, alpha6 integrin, and alpha-dystroglycan (alpha-DG). MATERIALS AND METHODS: Dissociated cells from 13.5-day postcoitum (dpc) fetal mouse pancreas were cultured for 4 days with laminin-1, with and without monoclonal antibodies and other agents known to block integrins or alpha-DG. Fetuses fixed in Bouin's solution or fetal pancreas cells fixed in 4% paraformaldehyde were processed for routine histology and for immunohistology to detect hormone expression and bromodeoxyuridine (BrdU) uptake. RESULTS: Blocking the binding of laminin-1 to alpha6 integrin with a monoclonal antibody, GoH3, abolished cell proliferation (BrdU uptake) and doubled the number of beta cells. Inhibition of molecules involved in alpha6 integrin signaling (phosphotidylinositol 3-kinase, F-actin, or mitogen-activated protein kinase) had a similar effect. Nevertheless, beta cells appeared to develop normally in alpha6 integrin-deficient fetuses. Blocking the binding of laminin-1 to alpha-DG with a monoclonal antibody, IIH6, dramatically decreased the number of beta cells. Heparin, also known to inhibit laminin-1 binding to alpha-DG, had a similar effect. In the presence of heparin, the increase in beta cells in response to blocking alpha integrin with GoH3 was abolished. CONCLUSIONS: These findings reveal an interplay between alpha6 integrin and alpha-DG to regulate laminin-1-induced beta-cell development. Laminin-I had a dominant effect via alpha-DG to promote cell survival and beta-cell differentiation, which was modestly inhibited by alpha6 signaling.

[Clinical observation of compound salvia injection in treating mid-severe infantile hypoxic-ischemic encephalopathy].

OBJECTIVE: To observe the changes of endothelin-1 (ET-1), nitrogen oxide (NO) and creatine phosphokinase BB isozyme (CK-BB) in blood and cerebrospinal fluid (CSF) in patients of infantile hypoxic-ishemic encephalopathy (HIE), and explore the efficacy of compound Salvia injection (CSI) in treating mid-severe HIE. METHODS: Sixty mid-severe infantile HIE patients were divided randomly into the treated and the control group. To the treated group CSI was added on the basis of conventional treatment, and to the control group the conventional treatment was given alone. The blood and CSF content of ET-1, NO and CK-BB at acute and convalescent stage in the two groups were determined and the therapeutic effects were compared between the two groups. RESULTS: The markedly effective rate and effective rate of the treated group was 80.0% and 93.3% respectively, while that of the control group was 66.7% and 83.3% respectively, the therapeutic effect in the treated group were obviously superior to that in the control group, the difference was significant (P < 0.05). CONCLUSION: ET-1, NO and CK-BB participated the pathological process of HIE. CSI was markedly effective in treating mid and severe HIE infants.

Male germ cell transplantation: promise and problems.

Male germ cell transplantation is a novel technique in which donor male stem germ cells are surgically transferred to the seminiferous tubules of a recipient testis by direct injection or via the rete testis or efferent duct. All germ cells that are destined to become stem spermatogonia are defined as male stem germ cells, including primordial germ cells from the gonadal ridges, and gonocytes and stem spermatogonia from the testis, all of which are transplantable and capable of undergoing normal spermatogenesis. Xenotransplantation of male germ cells from one species into the testis of another species, including human testicular cells in the mouse, has so far proved to be unsuccessful. However, the immunodeficient mouse testis can support rat spermatogenesis and produce apparently normal rat spermatozoa. The underlying mechanisms remain elusive. The present mini-review will focus on the importance of stem spermatogonial transplantation for testicular stem cell biology and discuss the likelihood of immune rejection after transplantation, which may limit the success of all male germ cell transplantation.

Laminin-1 promotes differentiation of fetal mouse pancreatic beta-cells.

Extracellular factors that regulate the growth and differentiation of cell lineages in the pancreatic primordia are poorly understood. Identification of these factors for pancreatic islet beta-cells could open new avenues for the treatment of insulin-dependent diabetes. We developed a low cell density serum-free culture system for dissociated pancreatic cells from the 13.5-day mouse fetus and investigated the effects of extracellular matrix proteins on differentiation of islet cells. After 4 days in culture, total cell number decreased by two-thirds, but insulin-positive beta-cell number increased 10-fold. Both of collagens I and IV inhibited cell survival (by >50%), whereas fibronectin had no effect. In the presence of soluble laminin-1, however, the number of beta-cells increased linearly by 60-fold without an increase in the total cell number; glucagon-positive cell number was unchanged, and somatostatin and pancreatic polypeptide-positive cells were not detected. The effect of laminin-1 was completely blocked by a monoclonal rat anti-laminin-1 antibody. In the presence of laminin-1, the thymidine analogue, BrdU, was incorporated into only 2.5% of cells, which were mainly insulin-negative at days 1-3. Laminin-1 appeared, therefore, to induce differentiation of beta-cells from precursor cells in day-13.5 fetal pancreas. Laminin-1 was shown to be expressed in the epithelial basement membrane of the 13.5- to 17.5-day fetal pancreas. These findings provide the first evidence of a role for laminin-1 to promote differentiation of pancreatic beta-cells.

Male germ cell transplantation: present achievements and future prospects.

Germ cells are unique, since their surviving descendants can undergo meiosis and differentiate into gametes, which transmit genetic material from one generation to another. We now know that male germ cells, whether they be primordial germ cells in gonadal ridges, gonocytes, or stem spermatogonia, are transplantable. The donor cells can be transferred by direct microinjection into the seminiferous tubules, rete testis or efferent ducts, depending on the recipient species. Following transplantation, the donor cells undergo spermatogenesis in the host's seminiferous tubules in rats and mice, and have even sired offspring in mice. Interspecific germ cell transfer is possible if the recipient's immune system is defective; nude or SCID mice can even produce rat spermatozoa. However, the major obstacle restricting widespread use of this new technology is its extremely low success rate. This article discusses some ideas for improving the success rate of the transfer technique, and considers several potential applications.

Behaviour of spermatogonia following recovery from busulfan treatment in the rat.

This study describes the morphological behaviour of spermatogonia following recovery from two doses of busulfan treatment in the rat. Twenty days after the second intraperitoneal injection of busulfan, the testes lost most of their spermatogenic cells and there were fewer dispersed singly surviving spermatogonia. These surviving cells were in close contact with the basal portions of adjacent Sertoli cells and the shrunken basal lamina, and were the source for repopulating the depleted seminiferous epithelium. During the initial stage of repopulation (48 days later), surviving spermatogonia underwent a phase of active proliferation: type A spermatogonia underwent symmetric and asymmetric divisions; type B spermatogonia underwent asynchronous differentiation. At day 96, normal spermatogenesis was fully recovered in many seminiferous tubules, represented by 80% of the rats regaining various degrees of fertility at day 120. These data provide an additional model for the study of self-renewal of stem spermatogonia and suggest that the asymmetric division of type A spermatogonia and their close contact with both the basal lamina and the Sertoli cells may be involved in regulating the number of stem spermatogonia and the delicate process of normal spermatogenesis.

Different fate of primordial germ cells and gonocytes following transplantation.

We have previously demonstrated that both donor primordial germ cells (PGCs) and gonocytes are capable of establishing spermatogenesis in the lumen of the seminiferous tubules of an adult host following transplantation in rats. Here we show that the PGCs, either in crude suspensions or after purification, undergo spermatogenesis only in the intraluminal compartment of the host's seminiferous tubules, while 4-5 days postpartum gonocytes also interdigitate with the host's seminiferous epithelium. The donor seminiferous epithelium was always in synchrony with the cycles of the host's spermatogenesis. It seems that the pattern of spermatogenesis of donor germ cells following transplantation in terms of its spacial location and the connection with the host's seminiferous epithelium depends on their developmental stages at transfer.

Purification and characterization of primordial germ cells in male rat fetuses.

The primordial germ cells in the testes of male rat fetuses at 15.5 days post coitum were purified by equilibrium centrifugation on a discontinuous Percoll gradient column after dissociation by trypsin-EDTA treatment and characterized before and after purification. Gonadal dimorphism first becomes evident at 14.5 days post coitum but the "stripey" appearance can not be observed in every fetal testis. Complete dissociation of the testes was essential for successful purification of the primordial germ cells. Using the isolation technique described here, over 50% of the primordial germ cells (around 10,000) could be harvested from each fetal testis with about 95% viability as determined by trypan blue exclusion and 90% purity, as determined by alkaline phosphatase histochemistry. Prior to and following purification, the primordial germ cells were characterized by their large size and round or oval nuclei. Their nucleoli were large and located centrally or eccentrically in the nucleus. The mitochondria were large, and round or oval, and there were a great number of ribosomes and polysomes dispersed in the cytoplasm. All these characteristics were similar to that of type A spermatogonia in postnatal testes. These morphological aspects correlate the observation that the primordial germ cells may be able to undergo normal spermatogenesis in an adult recipient testis.

Ultrastructural characteristics of primordial germ cells and their amoeboid movement to the gonadal ridges in the tammar wallaby.

The primordial germ cells (PGCs) of tammar wallaby fetuses are large cells with large nuclei. The cytoplasm of the PGCs contains characteristic spherical mitochondria and abundant ribosomes that make the cyoplasm appear dense. No permanent junctional complexes between PGCs and somatic cells were observed, and there were few cytoplasmic inclusions. The majority of migrating PGCs were observed outside the gonad in the dorsal mesentery and in the tissues adjacent to the gonad. The migrating PGCs have many finger-like, blunt pseudopodia, within which microfilament bundles are observed. The numerous dumbell-like PGCs with polarised cytoplasm suggest that the PGCs of this marsupial move to the gonadal ridges by amoeboid movement, but once the PGCs reach the gonadal ridge, they assume their more characteristic ovoid shape.

Male germ cell transplantation in rats: apparent synchronization of spermatogenesis between host and donor seminiferous epithelia.

Primordial germ cells (PGC) and gonocytes from male Sprague-Dawley rat fetuses and neonates were transplanted via the rete testis into the lumen of the seminiferous tubules of recipient adult Long Evans rats. The donor germ cells apparently differentiated into mini-tubules or irregular segments of seminiferous epithelium within the lumen of the host seminiferous tubules, and exhibited qualitatively normal spermatogenesis in 10 out of 16 recipients. The stage of spermatogenesis of the intraluminal epithelium was synchronized closely with that of the adjacent seminiferous tubule epithelium, suggesting that the spermatogenic cycle is regulated locally by the intraluminal microenvironment. Male germ cell transplantation provides an interesting new tool for investigating the control of spermatogenesis.

Postnatal differentiation and development of the rat epididymis: a stereological study.

Postnatal development and differentiation of the rat epididymis was studied in the rat from 15 to 120 days of life using stereological techniques. Both the relative volume (volume density) and absolute volume of the epithelial, interstitial, and luminal compartments in the initial segment, caput, corpus, and cauda epididymides were determined. In all segments the volume density of the epithelial compartment increased between days 15 and 30 before falling to adult values at 45 days in the initial segment (0.476 +/- 0.031), at 60 days in the caput (0.258 +/- 0.028) and at 90 days in the corpus (0.245 +/- 0.007) and cauda (0.140 +/- 0.004). The relative volume of the interstitium decreased, whilst that of the lumen increased over the same period with adult values being achieved earlier in the proximal segments than in the distal segments. In contrast to volume fraction the absolute volume of all compartments in all segments increased from day 15 to day 90. Between 90 and 120 days the absolute volumes of compartments in the initial segment and caput showed little volume change. All compartments in the corpus and cauda showed significant increases in volume over the same period. A similar pattern of development was observed with respect to the surface area of both the luminal and basement membrane aspects of the epithelium; surface area per unit volume (surface density) in all segments reached adult values at approximately 60 days, whilst the increase in absolute area of the surfaces ceased at 90 days in the initial segment and caput and continued to 120 days in the corpus and cauda.(ABSTRACT TRUNCATED AT 250 WORDS)