Proteomic analysis reveals alterations in the renal kallikrein pathway during hypoxia-induced hypertension

Obstructive sleep apnea syndrome (OSAS), a disorder characterized by episodic hypoxia (EH) during sleep, is associated with systemic hypertension. We used proteomic analysis to examine differences in rat kidney protein expression during EH, and their potential relationship to EH-induced hypertension. Young male Sprague-Dawley rats were exposed to either EH or sustained hypoxia (SH) for 14 (EH14/SH14) and 30 (EH30/SH30) days. Mean arterial blood pressure was significantly increased only in EH30 (p < 0.0002). Kidney proteins were resolved by two-dimensional-PAGE and were identified by MALDI-MS. Renal expression of kallistatin, a potent vasodilator, was down-regulated in all animals. Expression of alpha-1-antitrypsin, an inhibitor of kallikrein activation, was up-regulated in EH but down-regulated in SH. Western blotting showed significant elevation of B(2)-bradykinin receptor expression in all normotensive animals but remained unchanged in hypertensive animals. Proteins relevant to vascular hypertrophy, such as smooth muscle myosin and protein-disulfide isomerase were up-regulated in EH30 but were down-regulated in SH30. These data indicate that EH induces changes in renal protein expression consistent with impairment of vasodilation mediated by the kallikrein-kallistatin pathway and vascular hypertrophy. In contrast, SH-induced changes suggest the kallikrein- and bradykinin-mediated compensatory mechanisms for prevention of hypertension and vascular remodeling. To test the hypothesis suggested by the proteomic data, we measured the effect of EH on blood pressure in transgenic hKLK1 rats that overexpress human kallikrein. Transgenic hKLK1 animals were protected from EH-induced hypertension. We conclude that EH-induced hypertension may result, at least in part, from altered regulation of the renal kallikrein system.     

MAVG: locating non-overlapping maximum average segments in a given sequence

SUMMARY: MAVG is a software tool for finding k non-overlapping maximum-average segments that are sufficiently long in a given sequence of real numbers, for any k > 0. It has applications in several areas of biomolecular sequence analysis including locating GC-rich regions and CpG islands in a genomic sequence, and annotating multiple sequence alignments. AVAILABILITY: http://iubio.bio.indiana.edu/soft/molbio/pattern/cpg_islands/.     

Purification of industrial hydantoinase in one chromatographic step without affinity tag

Hydantoinase is used in industry as a biocatalyst for the production of optically pure D- or L-amino acids. Previously, homogeneous hydantoinase was obtained by multi-chromatographic purification procedures. Here, we reported a process that contained only a single chromatographic step to purify a recombinant hydantoinase to homogeneity. Hydantoinase from Agrobacterium radiobacter NRRL B11291 was expressed in Escherichia coli. The recombinant enzyme was purified following heat treatments, high concentration alcohol precipitation, and chelating Sephacel chromatography. The recombinant hydantoinase did not contain any affinity tags from the plasmid. This simplified procedure provided a convenient way to obtain hydantoinase in high yield (71%) and high purity. It should be very useful for further industrial application and for the study of the structure-function of hydantoinase.     

Effect of synthetic oligopeptides on osteoporosis

The objective of this study was to establish the solution method of GHRPS, the synthetic oligopeptides Tyr-Gly-Gly-Phe-Met-NH2, Tyr-Gly-Gly-Phe-Met-OH, Tyr-Gly-Gly-Phe-Leu-NH2, Tyr-Gly-Gly-Phe-Leu-OH, Tyr-Gly-Gly-Phe-Gly-NH2, and Tyr-Gly-Gly-Phe-Gly-OH, to verify their effect on osteoporosis. Male ICR mice (20 +/- 2 g) were used. The intramuscular injection dose of 6.3 mg/kg prednisone induced a significant decrease of body and femur weight of the animals. The subcutaneous injection dose of 18 microg/kg synthetic peptide was not effective to prevent the decrease of body and femur weight of the animals. The subcutaneous injection dose of 6.3 mg/kg prednisone elicited a decrease in content of femur calcium and in the level of serum calcium of the animals. The subcutaneous injection dose of 18 microg/kg Tyr-Gly-Gly-Phe-Leu-NH2, or Tyr-Gly-Gly-Phe-Leu-OH, or Tyr-Gly-Gly-Phe-Gly-NH2, significantly increased the content of femur calcium and decreased the level of serum calcium of the animals. It was also observed that the subcutaneous injection dose of 18 microg/kg Tyr-Gly-Gly-Phe-Gly-OH, Tyr-Gly-Gly-Phe-Leu-OH, Tyr-Gly-Gly-Phe-Met-OH, Tyr-Gly-Gly-Phe-Met-NH2 significantly increased the content of femur phosphorous and decreased the activity of ALP of the animals.     

Water-soluble degradable hyperbranched polyesters: novel candidates for drug delivery?

A novel approach to hyperbranched polymers is presented in this work. Hyperbranched polyesters with a large amount of terminal hydroxyl groups are prepared by a one-pot synthesis from commercially available AB-type and CD(n)-type monomers (n >/= 2). In this paper, Michael addition of diethanolamine (CD(2)) or N-methyl-d-glucamine (CD(5)) to methyl acrylate (AB) generates dominantly AD(n)-type intermediates. Further self-condensation of intermediates at higher temperature and in the presence of catalyst gives hyperbranched polyesters. Because of the tertiary amino groups in the backbone and the hydroxyl groups in the linear and terminal units, the resulting hyperbranched polyester is highly soluble in water. Furthermore, the hyperbranched polymer is degradable because of its ester units. So, the water-soluble hyperbranched polyesters might be applied as a novel material for drug delivery.     

A novel human hydroxysteroid dehydrogenase like 1 gene (HSDL1) is highly expressed in reproductive tissuesa

We report the cloning and characterization of a novel human hydroxysteroid dehydrogenase like gene (HSDL1) located on human chromosome 16q24.2. The HSDL1 cDNA is 3407 base pair in length, encoding a 309 amino acid polypeptide related to human 17-HSD3. Northern blot reveals that the HSDL1 is highly expressed in testis and ovary. In situ hybridization indicates that the expression of HSDL1 is predominantly increased in the prostate cancer tissue compared with the normal prostate tissue, which suggests that the gene expression is important to the arising of prostate cancer.     

Establishment and characterization of a cell line (HCDB-1) derived from a hamster buccal pouch carcinoma induced by DMBA and Taiwanese betel quid extract

This study identified that the carcinogenesis of hamster buccal pouch (HBP) induced by 7,12-dimethylbenz[a]anthracene (DMBA) was greatly enhanced (18 folds) by a combination treatment with Taiwanese betel quid (BQ) extract. A new cell line, HCDB-1, has been established from induced carcinomas. The cultured monolayer cells were epithelioid in shape with irregular nuclei. They demonstrated abundant cytokeratin and tonofilaments; however, ultrastructural well-organized desmosomes were lacking. The HCDB-1 cell exhibited population doubling in 19 h and was highly tumorigenic in nude mice. A C-->T transition at codon 141 (Ala to Val) of the p53 gene was detected in this cell. This mutation is equivalent to a specific temperature-sensitive mouse p53Ala135Val mutant that causes transformation by shifting to 37.5 degrees C. HCDB-1 is the first cell line established from the HBP model of oral carcinogenesis induced by DMBA/Taiwanese BQ extract. It might be valuable for exploring the molecular pathogenesis of oral cancer.     

Craniofacial shortening by contraction osteogenesis: an experimental model

Application of gradual external forces to correct craniofacial deformities challenges many procedures in conventional craniomaxillofacial surgery. Distraction osteogenesis is replacing traditional osteotomies for correction of patients with craniomaxillofacial deficiencies. However, the reverse concept, contraction osteogenesis, has yet to be established for patients with craniomaxillofacial excesses. The purpose of this investigation is to demonstrate the contraction osteogenesis phenomenon applied in a controlled animal model during the craniofacial growth period. Twenty-six 26-day-old rabbits were assigned to one of four groups: 0, control; 1, pin control (pin insertion); 2, no contraction (pins and contraction device application, without active contraction); and 3, contraction (pin insertion, contraction device application, and active contraction). An external fixator was placed across the incisive-maxillary suture, and the effects after 4.5 weeks of contraction at a rate of 0.5 mm twice a week were compared with control groups. The results were assessed by craniometric and cephalometric measurements and by histologic examination. Gross alterations were evident in the contraction group, characterized by midface anteroposterior shortening, maxillary regression, snout deviation, and anterior crossbite. Histologic examination of the contraction group demonstrated a significant increase in osteoblastic activity. Contraction osteogenesis is a new treatment concept in craniofacial development and may offer therapeutic opportunities for shortening skeletal structures without the need of osteotomies, thus taking advantage of the potential of craniofacial growth and remodeling.     

神经营养因子GDNF编码顺序的克隆及表达产物的纯化

根据基因库中的顺序,设计了胶质细胞源神经营养因子（ＧＤＮＦ）基因的ＰＣＲ引物,以此从人基因组ＤＮＡ中扩增并克隆了ＧＤＮＦ的编码序列,经ＤＮＡ测序确认后,该片段克隆到表达质粒ｐＥＴ－３ａ中,转化大肠杆菌ＢＬ２１（ＤＥ３）．培养的重组菌经ＩＰＴＧ诱导,在Ｔ７启动子调控下表达出ｈＧＤＮＦ蛋白．经电泳分析表明ＧＤＮＦ主要存在于细菌包涵体中．从培养菌中制备包涵体,经充分洗涤,溶解于含８ｍｏｌ／Ｌ尿素的变性缓冲液中．经ＳＰ－Ｓｅｐｈａｒｏｓｅ柱层析分离,梯度洗脱,以１５％ＳＤＳ－ＰＡＧＥ检查含ＧＤＮＦ的部分．将含单体ＧＤＮＦ部分进行复性,再次用ＳＰ－Ｓｅｐｈａｒｏｓｅ离子柱分离同源二体ＧＤＮＦ．最后经ＳＤＳ－ＰＡＧＥ制备电泳纯化,纯度大于９５％．经Ｎ端测序表明序列正确．经测定,每升培养菌可得约１０ｍｇ纯化的ＧＤＮＦ．     

Isothermal amplification detection of nucleic acids by a double-nicked beacon

Isothermal and rapid amplification detection of nucleic acids is an important technology in environmental monitoring, foodborne pathogen detection, and point-of-care clinical diagnostics. Here we have developed a novel method of isothermal signal amplification for single-stranded DNA (ssDNA) detection. The ssDNA target could be used as an initiator, coupled with a double-nicked molecular beacon, to originate amplification cycles, achieving cascade signal amplification. In addition, the method showed good specificity and strong anti-jamming capability. Overall, it is a one-pot and isothermal strand displacement amplification method without the requirement of a stepwise procedure, which greatly simplifies the experimental procedure and decreases the probability of contamination of samples. With its advantages, the method would be very useful to detect nucleic acids in point-of-care or field use.