The 3/4- and 3/6-Subfamily Variants of α-Conotoxins GI and MI Exhibit Potent Inhibitory Activity against Muscular Nicotinic Acetylcholine Receptors.

α-Conotoxins GI and MI belong to the 3/5 subfamily of α-conotoxins and potently inhibit muscular nicotinic acetylcholine receptors (nAChRs). To date, no 3/4- or 3/6-subfamily α-conotoxins have been reported to inhibit muscular nAChRs. In the present study, a series of new 3/4-, 3/6-, and 3/7-subfamily GI and MI variants were synthesized and functionally characterized by modifications of loop2. The results show that the 3/4-subfamily GI variant GI[∆8G]-II and the 3/6-subfamily variants GI[+13A], GI[+13R], and GI[+13K] displayed potent inhibition of muscular nAChRs expressed in Xenopus oocytes, with an IC50 of 45.4-73.4 nM, similar to or slightly lower than that of wild-type GI (42.0 nM). The toxicity of these GI variants in mice appeared to be about a half to a quarter of that of wild-type GI. At the same time, the 3/7-subfamily GI variants showed significantly lower in vitro potency and toxicity. On the other hand, similar to the 3/6-subfamily GI variants, the 3/6-subfamily MI variants MI[+14R] and MI[+14K] were also active after the addition of a basic amino acid, Arg or Lys, in loop2, but the activity was not maintained for the 3/4-subfamily MI variant MI[∆9G]. Interestingly, the disulfide bond connectivity "C1-C4, C2-C3" in the 3/4-subfamily variant GI[∆8G]-II was significantly more potent than the "C1-C3, C2-C4" connectivity found in wild-type GI and MI, suggesting that disulfide bond connectivity is easily affected in the rigid 3/4-subfamily α-conotoxins and that the disulfide bonds significantly impact the variants' function. This work is the first to demonstrate that 3/4- and 3/6-subfamily α-conotoxins potently inhibit muscular nAChRs, expanding our knowledge of α-conotoxins and providing new motifs for their further modifications.

Enhancing Mechanical Properties of Flexible Graphene/ Cellulose Conductive Paper by Chemically Modifying Cellulose Fibers.

In recent years, conductive paper made of graphene and cellulose fibers has triggered a great deal of attention due to its scalable fabrication process, low cost, and environmentally friendliness. However, poor mechanical properties limit its commercial applications. In this study, conductive graphene/cellulose paper with improved elastic modulus and desirable conductivity was achieved by chemically modifying the surface properties of natural cellulose fibers using aqueous 2,2,6,6-tetramethylpiperidine-1-oxyl (TEMPO) oxidation system. The effect of the carboxyl content on the mechanical properties and conductivity of graphene/cellulose paper was studied in detail. The results showed that the carboxyl content of cellulose had a significant impact on the mechanical properties of graphene/cellulose paper. At low carboxyl content (0.42 mmol/g), the elastic modulus of the graphene/oxidized-cellulose conductive paper achieved the value of 1572 MPa, which was 27.4% larger than that of cellulose/graphene conductive paper. Meanwhile, the as-prepared paper exhibited excellent conductive properties, and its sheet resistance was only 78.57 Ω/sq.

Expression of adrenomedullin and its receptor in lungs of rats with hypoxic pulmonary hypertension.

OBJECTIVE: To investigate the role of adrenomedullin (AM) in the development of hypoxic pulmonary hypertension (HPH), and to assess the expression of AM and adrenomedullin receptor (AMR) in the lungs of rats with HPH. METHODS: We exposed 10 rats to normobaric hypoxic conditions for 3 weeks to establish rat model of pulmonary hypertension; and 10 other rats were used as normoxic controls. Mean pulmonary arterial pressure (mPAP) was measured by a right cardiac catheterization. The thickness of pulmonary arterioles was measured by a computerized image analyzer. We used the reverse transcription-polymerase chain reaction (RT-PCR) to assess the change of expression of AM and AMR in lung of HPH rat model. RESULTS: Compared with the control group, hypoxic rats developed remarkable pulmonary hypertension, increment in the thickness of pulmonary arterioles and right ventricular hypertrophy (P < 0.01). Chronic hypoxia elicited a considerable increment in expression of AM and AMR in the lungs of rats, and the ratio of AM/beta-actin and AMR/beta-actin in lungs of rats treated with hypoxia were significantly higher (P < 0.01). CONCLUSIONS: The AM plays an important role in regulating pulmonary vascular tone and can ameliorate the development of hypoxic pulmonary hypertension in rats.

[Effect of adrenomedullin on the regulation of pulmonary arterial pressure in hypoxic rats].

OBJECTIVE: To evaluate the effect of adrenomedullin (ADM) on pulmonary circulation and the change of ADM in plasma and lung tissue from rats with hypoxic pulmonary hypertension. METHODS: Fifty Wistar rats were divided into the control group (10 rats) and the hypoxic group (40 rats). The animal model of pulmonary hypertension was established by exposing the rats to normobaric hypoxic conditions for 3 weeks; Mean pulmonary arterial pressure (mPAP) was measured by right cardiac catheterization, and mean systemic blood pressure (mSBP) was measured by left femoral catheterization. The thickness of pulmonary arterioles was measured by a computerized image analyser. The level of ADM in plasma and lung tissue was measured by radioimmunoassay. We ADM was administered in doses of 0.5, 1.0, 2.0 nmol/kg respectively to 30 hypoxic rats and the changes in mPAP and mSBP to ADM was evaluated. RESULTS: Rats exposed to hypoxia developed pulmonary hypertension, the mPAP in the control group being (16 +/- 3) mm Hg and in hypoxic group being (30 +/- 4) mm Hg, and the difference was significant (P < 0.01). The hypoxic rats developed significantly thickened pulmonary arterioles. The plasma level of ADM was (288 +/- 24) pg/ml in the hypoxic rats and (168 +/- 25) pg/ml in the control group the difference being significant (P < 0.01). The level of ADM in the lung homogenates from the hypoxic group was (2 319 +/- 238) pg/g and that from the control group was (1 153 +/- 127) pg/g and the difference was significant (P < 0.01). The ADM levels had a positive correlation with the mPAP (gamma = 0.567 and 0.612 P < 0.01, respectively). Administration of exogenous ADM reduced the mPAP in a dose-dependent manner in hypoxic rats, and the effect lasted 5 approximately 15 minutes. CONCLUSION: ADM has a relaxing effect on pulmonary circulation. The change of ADM in plasma and lung tissue may serve as a compensatory mechanism in maintaining the stability of pulmonary circulation in hypoxic condition.

[Effect of dehydroepiandrosterone on Ca(2+)-activated K+ channel of pulmonary arterial smooth muscle cells in pulmonary hypertensive rats].

OBJECTIVE: To investigate the effects of dehydroepiandrosterone (DHEA) on Ca(2+)-activated K(+) (K(Ca)) channel and mean pulmonary arterial pressure (mPAP) in rats with chronic pulmonary hypertension. METHODS: Fifty Wistar rats were divided randomly into a normal group (group A, n = 10) and a chronic hypoxia group (group B, n = 40). The rats in group B were subdivided into group B(1), B(2), B(3), and B(4) (each n = 10) at random. The rats in group B were exposed to hypoxia (FiO(2) = 0.10 +/- 0.05) for 3 weeks, whereas the rats in group A maintained in air. Under normoxic conditions, the smooth muscle cells (SMCs) were isolated from the pulmonary artery by the acute enzymatic dissection methods. In the symmetrical high K(+) solution, the K(Ca) currents were separated with inside-out configuration using the patch clamp technique. The activity of K(Ca) currents in SMCs between group B(1) and group A was compared under normoxic conditions, and the effect of DHEA on K(Ca) channel from group B(1) was observed. The mPAP and mean systemic arterial pressure (mSAP) were determined by right cardiac catheterization in rats from group B(2), B(3), B(4) before and after DHEA was administrated to rats by intravenous injection. RESULTS: The activity of K(Ca) channel in group B rats was much lower than that in group A (P < 0.01). DHEA could reverse the reduced K(Ca) channel in group B(1) rats. The mPAP were decreased significantly (P < 0.01) after DHEA was administrated to the rats in group B(2), B(3), B(4) with little change on mSAP (P < 0.05). CONCLUSIONS: Persistent decrease of K(Ca) channel activity may take part in the development of chronic hypoxic pulmonary hypertension in rats. DHEA can decrease the increased mPAP induced by chronic hypoxia via activating K(Ca) channel of SMCs from pulmonary arteries.

[Effect of levcromakalim and cromakalim on ATP-sensitive K+ channel of pulmonary arterial smooth muscle cells in pulmonary hypertensive rats].

OBJECTIVE: To investigate the effect of levcromakalim and cromakalim on ATP-sensitive K(+) channel (K(ATP)) and mean pulmonary arterial pressure (mPAP) in rats with chronic hypoxic pulmonary hypertension. METHODS: Ninety Wistar rats were divided randomly into a normal group (15) and a chronic hypoxia group (75). Rats in the hypoxia group were exposed to hypoxia (FiO(2) = 0.10 +/- 0.05) for 3 weeks, whereas rats in the normal group maintained in air. The currents of K(ATP) of pulmonary artery smooth muscle cells (PASMCs) from both the chronic hypoxic and the normal groups were observed using patch-clamp technique inside-out configuration, and the effect of levcromakalim and cromakalim on K(ATP) from chronic hypoxic group was determined under normoxic conditions. The mPAP and mSAP were determined by catheterization in hypoxic rats before and after levcromakalim and cromakalim were administrated respectively to the rats with doses ranging from 100 micro g/kg to 200 micro g/kg intravenously. RESULTS: Chronic hypoxia had little effect on the activity of K(ATP) in the chronic hypoxic rats. Levcromakalim and cromakalim remarkably activated the K(ATP) in chronic hypoxic rats. A dose-dependent decreasing effect on mPAP was elicited by levcromakalim and cromakalim in the hypoxic rats. CONCLUSION: Although K(ATP) activity is probably not directly involved in the development of chronic hypoxic pulmonary hypertension in rats, both levcromakalim and cromakalim decrease the increased mPAP induced by chronic hypoxia by activating K(ATP) of SMCs from pulmonary arteries, therefore diminishing the inhibitory effect of hypoxia on other K(+) channels.

Folate intake, methylenetetrahydrofolate reductase polymorphisms in association with the prognosis of esophageal squamous cell carcinoma.

AIM: An epidemiological study was conducted based on an esophageal cancer patient's cohort to investigate the association of folate intake and MTHFR C677T polymorphism with the prognosis of esophageal cancer in a Chinese population. METHODS: 167 patients aged 37-75 years who had histological confirmed diagnosis of esophageal squamous cell cancer were collected from Jan. 2006 to Jan. 2008. MTHFR genotypes at the C677T site were analyzed by PCR-based RFLP methods, and the folate intake was computed by multiplying the food intake (in grams) and the folate content (per gram) of food in our questionnaire. RESULTS: We found associations between the prognosis of esophageal cancer and smoking status, T and N stages. Individuals carrying the MTHFR 677CT and TT genotypes showed a shorter survival time than with the CC genotype, with adjusted HRs (95% CI) of 1.20 (0.56-2.15) and 2.29 (1.30-4.28), respectively. Similarly, those carrying MTHFR 677T allele had a 1.86-fold risk of death. A higher folate concentration showed a significant decreased risk of death, with an HR (95% CI) of 0.45 (0.18-0.87). Individuals with high folate intake and the MTHFR 677CC genotype showed a significant decreased risk of esophageal cancer (0.43, 0.25-0.89). CONCLUSION: Our findings supports the hypothesis that high folate intake and active MTHFR C677T polymorphism may exert protective roles in the prognosis of esophageal cancer in the Chinese population.

Aberrant DNA methylation of P16, MGMT, and hMLH1 genes in combination with MTHFR C677T genetic polymorphism and folate intake in esophageal squamous cell carcinoma.

AIM: The present case-control study was conducted to explore the association of MTHFR gene polymorphism and relations of P16, MGMT and HMLH1 to MTHFR and folate intake. METHODS: A total of 257 cases of esophageal squamous cell carcinoma confirmed by histopathological examination were collected. Genotyping of P16, MGMT and HMLH1 was accomplished by methylation-specific polymerase chain reaction (PCR) after sodium bisulfate modification of DNA and the MTHFR C677T genetic polymorphism was detected by PCR- restriction fragment-length polymorphism (PCR-RFLP). RESULTS: The proportions of DNA hypermethylation in P16, MGMT and hMLH1 in cancer tissues were significantly higher than in paracancerous normal tissue. The proportion of hypermethylation in at least one gene was 88.5% in cancer tissue, and was also significantly higher than that in paracancerous normal tissue. Our finding showed individuals with homozygotes (TT) of MTHFR C677T had significant risk of DNA hypermethylation of MGMT in cancer tissues, with an OR (95% CI) of 3.15 (1.12-6.87). Similarly, patients with high intake of folate also showed a slight high risk of DNA methylation of MGMT, with OR (95% CI) of 2.03 (1.05-4.57). CONCLUSION: Our study found the P16, MGMT and hMLH1 demonstrate a high proportion of hypermethylation in esophageal squamous cell cancer cancer tissues, which might be used as biomarkers for cancer detection.

Glulathione-S-transferases gene polymorphism in prediction of gastric cancer risk by smoking and Helicobacter pylori infection status.

AIM: To evaluate the association of glutathione S-transferases gene polymorphisms with the risk of gastric cancer, with reference to smoking and Helicobacter pylori infection. METHODS: We conducted a 1:1 matched case-control study with 410 gastric cancer cases and 410 cancer-free controls. Polymorphisms of GSTM1, GSTT1 and GSTP1 were determined using PCR-CTPP. RESULTS: The GSTM1 and GSTT1 null genotypes were significantly associated with the risk of gastric cancer after adjusting for potential confounding factors (OR=1.68, 95% CI=1.32-2.23 for null GSTM1, OR=1.73; 95% CI=1.24-2.13 for null GSTT1). The combination of null GSTM1 and null GSTT1 conferred an elevated risk (OR=2.54, 95% CI=1.55-3.39). However, no association was found for GSTP1 polymorphism The smoking modified the association of GSTM1 and GSTT1 null genotypes with the risk of gastric cancer. CONCLUSION: GSTM1 and GSTT1 null genotypes are associated with increased risk of gastric cancer, and smoking modifies the association.