We purified from bovine serum a glycoprotein that promotes the survival of rat embryonic neurons cultured from septum and other brain regions. A 40,000-fold purification was achieved by using a combination of ammonium sulfate precipitation, Zn2+ affinity chromatography, Cibacron blue 3-GA dye affinity chromatography, ABx ion exchange chromatography, and preparative PAGE. The active protein had an apparent molecular weight of 50-60 kDa. The concentration required for half-maximal survival (EC50) was 12 ng/ml ( approximately 200 pM) for the final fraction. Amino acid sequencing after cyanogen bromide cleavage yielded two sequences that are homologous to regions of deduced sequence of the selenoprotein-P (SPP) family in bovine, rat, and human. Antibodies against a synthetic peptide within the bovine SPP sequence immunoprecipitated and inhibited the survival-promoting activity of a partially purified serum fraction. The purified protein supported neuronal survival more effectively than inorganic selenium. These results suggest that SPP or an SPP-like protein contributes to the neuronal survival-promoting activity of serum.

No abstract

Pulmonary arterial hypertension (PAH) is the most common form of pulmonary hypertension. Pulmonary arterial remodeling is closely related to the abnormal proliferation of pulmonary artery smooth muscle cells (PASMCs), which leads to the thickening of the medial layer of muscular arteries and then results in the narrowing or occlusion of the precapillary arterioles and PAH. However, the mechanisms underlying the abnormal proliferation of PASMCs remain unclear. In this study, we established rat primary PAH models using monocrotaline (MCT) injection or hypoxic exposure, then investigated the expression patterns of seven miRNAs associated with multiple pathogenic pathways central to pulmonary hypertension, and further explored the roles and the possible mechanisms of miR-135a during the development of PAH. In the rat primary PAH models, we observed that the expression of miR-135a-5p in lungs was drastically decreased at the initial stage of PAH development after MCT administration or hypoxic exposure, but it increased by 12-fold or 10-fold at the later stage. In vitro study in PASMCs showed a similar pattern of miR-135a-5p expression, with downregulation at 6 h but upregulation at 18, 24, and 48 h after hypoxic exposure. Early, but not late, administration of a miR-135a-5p mimic inhibited hypoxia-induced proliferation of PASMCs. The protective role of early miR-135a-5p agomir in the PAH rat model further supported the hypothesis that the early decrease in the expression of miR-135a-5p contributes to the proliferation of PASMCs and development of PAH, as early administration of miR-135a-5p agomir (10 nM, i.v.) reversed the elevated mean pulmonary arterial pressure and pulmonary vascular remodeling in MCT-treated rats. We revealed that miR-135a-5p directly bound to the 3'-UTR sequence of rat transient receptor potential channel 1 (TRPC1) mRNA and decreased TRPC1 protein expression, thus inhibiting PASMC proliferation. Collectively, our data suggest that dysregulation of miR-135a-5p in PASMCs contributes to the abnormal proliferation of PASMCs and the pathogenesis of PAH. Increasing miR-135a-5p expression at the early stage of PAH is a potential new avenue to prevent PAH development.

Hydrogen sulfide (H2S), is a member of the novel family of endogenous gaseous transmitters, termed “gasotransmitters exhibiting diverse physiological activities, and is generated in mammalian tissues mainly by cystathionine β-synthase (CBS), cystathionine γ-lyase (CSE) and 3-mercaptopyruvate sulfurtransferase (3MST) in conjunction with cysteine (aspartate) aminotranferase (CAT). The distributions of these enzymes are species- and tissue-specific. The liver, as the main organ that generates H2S in vivo, functions in biotransformation and metabolism. However, the liver is vulnerable to damage from internal and external factors, including inflammatory mediators, drugs and poisons. The present study evaluated the endogenous CBS-H2S synthesis regulating lipopolysaccharide (LPS)-induced apoptosis of hepatic cells. The rat hepatic cell line, BRL, was incubated with LPS for various time periods to establish a cell-damage model. Incubation with LPS resulted in a significant increase in CBS expression and H2S production. It also stimulated apoptosis and decreased the mitochondrial membrane potential. Pretreatment with the CBS inhibitor aminooxyacetic acid (AOAA) or CBS small interfering RNA (siRNA) decreased LPS-enhanced H2S production. Notably, apoptosis increased for a short period and then decreased gradually, while the mitochondrial membrane potential demonstrated the opposite trend. These results showed that endogenous CBS-H2S synthesis demonstrated early anti-apoptotic activity and subsequent pro-apoptotic activity in LPS-induced apoptosis. These results suggest a new approach for developing novel drugs for this condition.