Internalization of insulin and its receptor in the isolated rat adipose cell. Time-course and insulin concentration dependency

The time-course and insulin concentration dependency of internalization of insulin and its receptor have been examined in isolated rat adipose cells at 37 degrees C. The internalization of insulin was assessed by examining the subcellular distribution of cell-associated [125I]insulin among plasma membrane, and high-density (endoplasmic reticulum-enriched) and low-density (Golgi-enriched) microsomal membrane fractions prepared by differential ultracentrifugation. The distribution of receptors was measured by the steady-state exchange binding of fresh [125I]insulin to these same membrane fractions. At 37 degrees C, insulin binding to intact cells is accompanied initially by the rapid appearance of intact insulin in the plasma membrane fraction, and subsequently, by its rapid appearance in both the high-density and low-density microsomal membrane fractions. An apparent steady-state distribution of insulin per mg of membrane protein among these subcellular fractions is achieved within 30 min in a ratio of 1:1.54:0.80, respectively. Concomitantly, insulin binding to intact cells is associated with the rapid disappearance of approx. 30% of the insulin receptors initially present in the plasma membrane fraction and appearance of 20-30% of those lost in the low-density microsomal membrane fraction. However, the number of receptors in the high-density microsomal membrane fraction does not change. This redistribution of receptors also appears to reach a steady-state within 30 min. Both processes are insulin concentration-dependent, correlating with receptor occupancy in the intact cell, and are partially inhibited at 16 degrees C. While the steady-state subcellular distributions of insulin and its receptor do not correlate with that of acid phosphatase, chloroquine markedly increases the levels of insulin associated with all three membrane fractions in apparent proportion to the distribution of this lysosomal marker enzyme activity, without more than marginally potentiating insulin's effects on the distribution of receptors. These results demonstrate that insulin, initially bound to the plasma membrane of the isolated rat adipose cell, is rapidly translocated by a receptor-mediated process into at least two intracellular compartments associated with the cell's high- and low-density microsomes. Furthermore, insulin simultaneously induces the translocation of its own receptor from the plasma membrane into the latter compartment. These translocations appear to represent the internalization and partial dissociation of the insulin-receptor complex through insulin-induced receptor cycling.     

A thrombin receptor in resident rat peritoneal macrophages.

Resident rat peritoneal macrophages possess 6 x 10(2) high-affinity binding sites per cell for bovine thrombin with a Kd of 11 pM, and 7.5 x 10(4) low-affinity sites with a Kd of 5.8 nM. These binding sites are highly specific for thrombin. Half-maximal binding of 125I-labeled bovine thrombin is achieved after 1 min at 37 degrees C, and after 12 min at 4 degrees C. The reversibly bound fraction of the ligand dissociates according to a biexponential time course with the rate constants 0.27 and 0.06 min-1 at 4 degrees C. Part of the tracer remains cell-associated even after prolonged incubation, but all cell-associated radio-activity migrates as intact thrombin upon sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The bound thrombin is minimally endocytosed as judged by the resistance to pH 3 treatment, and the receptor does not mediate a quantitatively important degradation of the ligand. The binding is not dependent on the catalytic site of thrombin, since irreversibly inactivated thrombin also binds to the receptor. 125I-labeled thrombin covalently cross-linked to its receptor migrates in sodium dodecyl sulfate-polyacrylamide gel electrophoresis with a Mr 160,000, corresponding to an approximate receptor size of Mr 120,000.     

抑菌生的研究总结报告

本文报告一种由枯草杆菌制成的生态制剂,并命名为抑菌生(Subtilobiogen)。该制剂对创、烧伤感染有治疗作用。经过安全试验、急性毒性试验、Ames试验和微核试验证明,该制剂是一种无害、无毒和有致突变作用的活菌制剂。抑菌生有膏剂、乳剂及粉剂3种剂型。抑菌生在试管和体内对金黄色葡萄球菌、绿脓杆菌和大肠杆菌均具有抑菌作用。临床观察证明,将抑菌生喷洒在创面上,对浅Ⅱ°度、深Ⅱ°度及混合型烧伤感染均具有明显疗效。实验组(181例)与对照组(174例)相比较,在统计学上具有显著性差异。抑菌生的作用机制,经过初步试验证明,与营养争夺和占位性保护有关,因为枯草杆菌的生长速度超过金黄色葡萄球菌、绿脓杆菌和大肠杆菌的生长速度。     

Assessment of alcohol percentage test for fungal surface hydrophobicity measurement

Aim: To determine whether assessing the penetration of solutions with different concentrations of ethanol (alcohol percentage test: APT) on fungal surfaces is effective in characterization of hydrophobicity on fungal surfaces. Methods and Results: APT and contact angle (CA) measurements were conducted on nine hydrophobic and two hydrophilic fungal strains from the phyla of Ascomycota, Basidiomycota and Zygomycota. There was a strong positive correlation (R² = 0·95) between the APT and CA measurements from eight of the nine hydrophobic stains (four pathogenic and mycotoxigenic Fusarium taxa, one melanosporaceous biotrophic taxon, Alternaria sp, Penicillium aurantiogriseum and Cladosporium cladosporioides). Hydrophilic control strains, Mortierella hyalina and Laccaria laccata, had CAs <90° and no measurable degree of hydrophobicity using the APT method. Conclusions: The APT method was effective in measuring the degree of hydrophobicity and can be conducted on different zones of fungal growth. Significance and Impact of the Study: Characterization of fungal surface hydrophobicity is important for understanding of its particular role and function in fungal morphogenesis and pathogenesis. APT is a simple method that can be utilized for fungal hydrophobicity measurements when CA cannot be measured because of obscured view from aerial mycelia growth.     

Bmp2 regulates Serpinb6b expression via cAMP/PKA/Wnt4 pathway during uterine decidualization

Serpinb6b is a novel member of Serpinb family and found in germ and somatic cells of mouse gonads, but its physiological function in uterine decidualization remains unclear. The present study revealed that abundant Serpinb6b was noted in decidual cells, and advanced the proliferation and differentiation of stromal cells, indicating a creative role of Serpinb6b in uterine decidualization. Further analysis found that Serpinb6b modulated the expression of Mmp2 and Mmp9. Meanwhile, Serpinb6b was identified as a target of Bmp2 regulation in stromal differentiation. Treatment with rBmp2 resulted in an accumulation of intracellular cAMP level whose function in this differentiation program was mediated by Serpinb6b. Addition of PKA inhibitor H89 impeded the Bmp2 induction of Serpinb6b, whereas 8‐Br‐cAMP rescued the defect of Serpinb6b expression elicited by Bmp2 knock‐down. Attenuation of Serpinb6b greatly reduced the induction of constitutive Wnt4 activation on stromal cell differentiation. By contrast, overexpression of Serpinb6b prevented this inhibition of differentiation process by Wnt4 siRNA. Moreover, blockage of Wnt4 abrogated the up‐regulation of cAMP on Serpinb6b. Collectively, Serpinb6b mediates uterine decidualization via Mmp2/9 in response to Bmp2/cAMP/PKA/Wnt4 pathway.