Puromycin selectively increases mdr1a expression in immortalized rat brain endothelial cell lines.

The blood-brain barrier (BBB) plays an important role in controlling the passage of molecules from blood to brain extracellular fluid. The multidrug efflux pump P-glycoprotein (P-gp) is highly expressed in the luminal membrane of brain endothelium and contributes to the formation of a functional barrier to lipid-soluble drugs such as anticancer agents. The mdr1a P-gp-encoding gene is exclusively expressed in the rodent BBB. Primary cultures of rat brain endothelial cells and GP8.3 cells showed a dramatic decrease in mdr1a mRNA level and some expression of mdr1b mRNA. GPNT cells, derived from GP8.3 cells after transfection with a puromycin resistance gene, were chronically treated with 5 microg/mL puromycin, a P-gp substrate. Compared with rat brain endothelial cells and GP8.3 cells, GPNT cells exhibited a very high level of expression of mdr1a mRNA together with a moderate level of mdr1b mRNA expression. Accordingly, P-gp expression and activity were strongly increased. When GP8.3 and puromycin-starved GPNT cells were treated with puromycin, mdr1a expression was selectively increased. High expression of mdr1a mRNA in GPNT cells may thus be related to the chronic treatment with puromycin. We conclude that GPNT cells may be used as a valuable rat in vitro model for studying the regulation of mdr1a expression at the BBB level.

Differential regulation of the TRH gene promoter by triiodothyronine and dexamethasone in pancreatic islets.

TRH was initially found in the hypothalamus and regulates TSH secretion. TRH is also produced by insulin-containing beta-cells. Endogenous TRH positively regulates glucagon secretion and attenuates pancreatic exocrine secretion. We have previously shown that triiodothyronine (T(3)) down-regulates pre-pro-TRH gene expression in vivo and in vitro. The present study was designed to determine the initial impact of T(3) on rat TRH gene promoter and to compare this effect with that of dexamethasone (Dex). Primary islet cells and neoplastic cells (HIT T-15 and RIN m5F) were transiently transfected with fragments of the 5'-flanking sequence of TRH fused to the luciferase reporter gene. The persistence of high TRH concentrations in fetal islets in culture, probably due to transactivating factors, allowed us to explore how T(3) and Dex regulate the TRH promoter activity in transfected cells and whether the hormone effect is dependent on the cell type considered. TRH gene promoter activity is inhibited by T(3) in primary but not neoplastic cells and stimulated by Dex in both primary and neoplastic cells of islets. These findings validate previous in vivo and in vitro studies and indicate the transcriptional impact of these hormones on TRH gene expression in the pancreatic islets.

Triiodothyronine down-regulates thyrotropin-releasing hormone (TRH) synthesis and decreases pTRH-(160-169) and insulin releases from fetal rat islets in culture.

TRH is localized with insulin in beta-cells. It is synthesized as a prohormone containing five copies of TRH and seven cryptic peptides, including pro (p)-TRH-(160-169). Thyroid hormone regulates the expression of several genes encoding peptide hormones. We found that circulating T3 concentrations were inversely correlated with TRH levels in two physiopathological situations. There are low concentrations of circulating thyroid hormone and very high concentrations of TRH and pTRH-(160-169) during development, and experimental hypothyroidism results in higher concentrations of prepro (pp)-TRH messenger RNA (mRNA) and TRH content in the adult rat pancreas than are present in the euthyroid pancreas. The present study was carried out to investigate the interaction between T3 and pancreatic TRH during the functional maturation of islets in culture and to validate the data obtained in vivo. T3 decreases ppTRH mRNA in islets in a dose-dependent manner. The primary impact of T3 on islet function may be mediated by ppTRH mRNA, as short term T3 treatment had no effect. Long term T3 treatment reduced the islet TRH content and the amounts of pTRH-(160-169) and insulin released. This secretory pattern and coordinated regulation of pTRH-(160-169) and insulin suggests that pTRH-(160-169) plays a specific role in the regulation of insulin secretion.

Thyroid status and the regulation of thyrotropin-releasing hormone synthesis in rat pancreatic islets: comparison with insulin regulation.

Thyrotropin-releasing hormone (TRH), originally characterized as the first hypothalamic hormone, is also synthesized in the insulin-containing cells. TRH stimulates the glucagon secretion and attenuates exocrine pancreatic secretions. We have previously reported, using whole pancreatic homogenates, that TRH content increased in hypothyroid rats, associated to a loss of TRH-degrading activity. The present study was undertaken on purified islets, an appropriate model to examine thyroid status-dependent regulation of two islet hormones, TRH and insulin. The islets from hypothyroid rat pancreas had increased TRH content (4x) without any change in insulin content. Likewise, the Northern blot analysis revealed that the steady-state concentrations of TRH mRNA increased (4x) while those of insulin remain unchanged. These data therefore suggest that TRH gene transcription is under the negative control of T3. This study also provides insight into islet response to impaired thyroid function.

A new biological contribution of cyclo(His-Pro) to the peripheral inhibition of pancreatic secretion.

The tripeptide pyro-Glu-His-Pro-NH2[thyrotropin-releasing hormone (TRH)] was isolated from the hypothalamus as a thyrotropin-releasing factor. It has a broad spectrum of central nervous system-mediated actions, including the stimulation of exocrine pancreatic secretion. TRH is also synthesized in the endocrine pancreas and found in the systemic circulation. Enzymatic degradation of TRH in vivo produces other bioactive peptides such as cyclo(His-Pro). Because of the short half-life of TRH and the stability of cyclo(His-Pro) in vivo, we postulated that at least part of the peripheral TRH effects on the exocrine pancreatic secretion may be attributed to cyclo(His-Pro), which has been shown to have other biological activities. This study determines in parallel the peripheral effects of TRH and cyclo(His-Pro) as well as the putative contribution of other TRH-related peptides on exocrine pancreatic secretion in rats. TRH and its metabolite cyclo(His-Pro) dose dependently inhibited 2-deoxy-D-glucose (2-DG)-stimulated pancreatic secretion. TRH and all the related peptides tested had no effect on the basal and cholecystokinin-stimulated amylase release from pancreatic acinar cells in vitro. These data indicate that cyclo(His-Pro) mimics the peripheral inhibitory effect of TRH on 2-DG-stimulated exocrine pancreatic secretion. This effect is not detected on isolated pancreatic acini. Our findings provide a new biological contribution for cyclo(His-Pro) with potential experimental and clinical applications.

[Pulmonary and hepatic aspergillomas in the deer]. / Plicní a jaterní aspergilomy srnci zvere.

Four solitary aspergillomas in deer are being described. Two were found in the lungs and two in the liver. Morphologically, they were bulky, local lesions filled with homogenous, fragile, necrotic tissue of conspicuously green color. Pulmonary aspergillomas communicated with the conducting bronchus having affected fibers. The necrotic tissue of aspergillomas was interwoven by diffuse abundant fibers of Aspergillus fumigatus.

[Outbreak of paraventricular and ventricular mucormycosis in fattening bulls]. / Hromadny výskyt paraventrikulární a ventrikulární mukormykózy u býkü ve výkrmu

There is a description of mass occurrence of nine cases of acute, fatal paraventricular and ventricular mucormycosis in fattened bulls at the age of about 18 months. In the reticulum, the disease was characterized by numerous ulcerations penetrating into the submucosa. In rumen the deposits were of hemorrhagic character penetrating through a thick muscular layer into the subserosa. Hemorrhagic peritonitis was within the range of those changes. Thromboses of arteries and veins prevailed in the microscopic picture. Inflammatory reparative changes were formed by neutrophile leucocytes. Mucoraceum fibres were demonstrated in pathological manifestations. In three cases, strains of Rhizopus cohnii and Absidia ramosa were cultivated simultaneously from the sub-ulcerous layers in the reticulum. A number of findings testify that the mass occurrence of mucormycosis followed the feeding of mouldy bakery wastes on the basis of acidosis.