Phosphodiesterase-5 Gln817 is critical for cGMP, vardenafil, or sildenafil affinity: its orientation impacts cGMP but not cAMP affinity

The side group of an invariant Gln in cGMP- and cAMP-specific phosphodiesterases (PDE) is held in different orientations by bonds with other amino acids and purportedly discriminates between guanine and adenine in cGMP and cAMP. In cGMP-specific PDE5, Gln(775) constrains the orientation of the invariant Gln(817) side chain, which forms bidentate bonds with 5'-GMP, vardenafil, sildenafil, and 3-isobutyl-1-methylxanthine (IBMX) (Sung, B. J., Hwang, K. Y., Jeon, Y. H., Lee, J. I., Heo, Y. S., Kim, J. H., Moon, J., Yoon, J. M., Hyun, Y. L., Kim, E., Eum, S. J., Park, S. Y., Lee, J. O., Lee, T. G., Ro, S., and Cho, J. M. (2003) Nature 425, 98-102; Huai, Q., Liu, Y., Francis, S. H., Corbin, J. D., and Ke, H. (2004) J. Biol. Chem. 279, 13095-13101; Zhang, K. Y., Card, G. L., Suzuki, Y., Artis, D. R., Fong, D., Gillette, S., Hsieh, D., Neiman, J., West, B. L., Zhang, C., Milburn, M. V., Kim, S. H., Schlessinger, J., and Bollag, G. (2004) Mol. Cell 15, 279-286). PDE5(Q817A) and PDE5(Q775A) were generated to test the hypotheses that Gln(817) is critical for cyclic nucleotide or inhibitor affinity and that Gln(775) immobilizes the Gln(817) side chain to provide cGMP/cAMP selectivity. Allosteric cGMP binding and the molecular mass of the mutant proteins were unchanged compared with PDE5(WT). For PDE5(Q817A), K(m) for cGMP or cAMP was weakened 60- or 2-fold, respectively. For PDE5(Q775A), K(m) for cGMP was weakened approximately 20-fold but was unchanged for cAMP. For PDE5(Q817A), vardenafil, sildenafil, and IBMX inhibitory potencies were weakened 610-, 48-, and 60-fold, respectively, indicating that Gln(817) is a major determinant of potency, especially for vardenafil, and that binding of vardenafil and sildenafil differs substantially. Sildenafil and vardenafil affinity were not significantly affected in PDE5(Q775A). It is concluded that Gln(817) is a positive determinant for PDE5 affinity for cGMP and several inhibitors; Gln(775), which perhaps restricts rotation of Gln(817) side chain, is critical for cGMP affinity but has no measurable effect on affinity for cAMP, sildenafil, or vardenafil.     

Association of TNF-308 G>A polymorphism located in tumor necrosis factor a with the risk of developing cervical cancer and results of pap smear

Tumor necrosis factor a (TNFa) is an inflammatory cytokine that plays a crucial role in the immune response and the progression of cervical lesions. There is a growing body of data evaluating the value of a genetic variant in the TNFa gene with the risk of developing cervical cancer. The aim of this study was to explore the association of a variant, TNF-308 G>A, residing in the TNFa gene with cervical cancer. A total of 91 women with cervical cancer and 161 women as the control group were recruited. DNA was extracted, and Taqman®-probes-based assay was used for genotyping. Our results showed that the minor allele frequency was 0.3 in total population, and the frequency of minor allele A was more in the case group compared with the control. The regression models in different genetic models also revealed that the allele A is a potential risk factor for the development of cervical cancer. In particular, in the dominant model, patients with AG and AA genotypes had a higher risk of developing cervical cancer with odds ratio (OR) of 2.75 (95% confidence interval [CI]: 1.57-4.83, <0.001) and OR of 7.27 (95%CI: 2.5-20.8, <0.001), compared with the GG genotype. Moreover, a similar outcome was obtained for smear test results. Our study demonstrated that TNF-308 G>A located on TNF-a was associated with the risk of cervical cancer, supporting further studies in a larger population and multicenter setting to show the value of emerging markers as risk stratification biomarkers in cervical cancer.     

Dysregulation of apoptotic signaling in cancer: molecular mechanisms and therapeutic opportunities

Apoptosis is a tightly regulated cell suicide program that plays an essential role in the maintenance of tissue homeostasis by eliminating unnecessary or harmful cells. Defects in this native defense mechanism promote malignant transformation and frequently confer chemoresistance to transformed cells. Indeed, the evasion of apoptosis has been recognized as a hallmark of cancer. Given that multiple mechanisms function at many levels to orchestrate the regulation of apoptosis, a multitude of opportunities for apoptotic dysregulation are present within the intricate signaling network of cell. Several of the molecular mechanisms by which cancer cells are protected from apoptosis have been elucidated. These advances have facilitated the development of novel apoptosis-inducing agents that have demonstrated single-agent activity against various types of cancers cells and/or sensitized resistant cancer cells to conventional cytotoxic therapies. Herein, we will highlight several of the central modes of apoptotic dysregulation found in cancer. We will also discuss several therapeutic strategies that aim to reestablish the apoptotic response, and thereby eradicate cancer cells, including those that demonstrate resistance to traditional therapies.     

Intravenous immunoglobulin G treatment increases live birth rate in women with recurrent miscarriage and modulates regulatory and exhausted regulatory T cells frequency and function

Exhausted T cells and regulatory T (Treg) cells have been recently proposed to be new risk factors for recurrent miscarriage (RM). Intravenous immunoglobulin G (IVIG) treatment reported to modulate various immune cells. In this study, the effects of IVIG on the frequency and function of exhausted T cells, exhausted Tregs, and Treg cells, as well as pregnancy outcome in women with unexplained RM (URM), were investigated. Ninety-four pregnant women with RM were enrolled. At the time of positive pregnancy, blood samples were drawn. Forty-four patients with URM were included as IVIG receiving treated group and received 400 mg/kg of IVIG and the rest fifty patients were considered as a control group and received no IVIG administration. IVIG was given intravenously every 4 weeks during 32 weeks of gestation. Blood samples of patients were collected after the latest administration. Exhausted T cells, exhausted Tregs, and Treg cells were evaluated pre- and posttreatment in both groups. IVIG induced a significant decrease in the frequency of exhausted Tregs population and function as well as a significant increase in Treg cells population, however, IVIG failed to affect population and the function of exhausted T cells. Pregnancy outcome was successful in IVIG treated women (86.3%) and were significantly different (P = 0.0006) in compared with the untreated URM subjects (42%). Therefore, employing of IVIG increases Treg cells and diminishes exhausted Tregs responses in RM patients with cellular immune anomalies throughout the pregnancy. Immunemodulatory effects of IVIG are probably associated with successful pregnancy outcome.     

Current progress in cancer immunotherapy based on natural killer cells

One of the most common diseases in the present era is cancer. The common treatment methods used to control cancer include surgery, chemotherapy, and radiotherapy. Despite progress in the treatment of cancers, there still is no definite therapeutic approach. Among the currently proposed strategies, immunotherapy is a new approach that can provide better outcomes compared with existing therapies. Employing natural killer (NK) cells is one of the means of immunotherapy. As innate lymphocytes, NK cells are capable of rapidly responding to cancer cells without being sensitized or restricted to the cognate antigen in advance, as compared to T cells that are tumor antigen-specific. Latest insights into the biology of NK cells have clarified the underlying molecular mechanisms of NK cell maturation and differentiation, as well as controlling their effector functions through the investigation of the ligands and receptors engaged in recognizing cancer cells by NK cells. Elucidating the fact that NK cells recognize cancer cells could similarly show the mechanism through which cancer cells possibly avoid NK cell-dependent immune surveillance. Additionally, the expectations for novel immunotherapies by targeting NK cells have increased through the latest clinical outcomes of T–cell-targeted cancer immunotherapy. For this emerging method, researchers are still attempting to develop protocols for conferring the best proliferation and expansion medium, activation pathways, utilization dosage, transferring methods, as well as reducing possible side effects in cancer therapy. This study reviews the NK cells, their proliferation and expansion methods, and their recent applications in cancer immunotherapy.     

Improved production of 3‐hydroxypropionaldehyde by complex formation with bisulfite during biotransformation of glycerol

3‐Hydroxypropionaldehyde (3HPA) is an important specialty chemical which can be produced from glycerol using resting cells of Lactobacillus reuteri. This biocatalytic route, however, suffers from substrate‐ and product‐mediated loss of enzyme activity within 2 h of biotransformation. In order to overcome the inhibitory effects of 3HPA, complex formation with sodium bisulfite was investigated, optimized and applied for in situ capture of the aldehyde during biotransformation of glycerol in a fed‐batch process. As a result, the activity of the cells was maintained for at least 18 h. The 3HPA produced per gram cell dry weight was increased 5.7 times compared to the batch production process, and 2.2 times compared to fed‐batch process without in situ complex formation. This approach may have potential for production and in situ removal of 3HPA after further process development. Biotechnol. Bioeng. 2013; 110: 1243–1248. © 2012 Wiley Periodicals, Inc.     

Effect of temperature and ionic strength on structure and chaperone activity of glycated and non-glycated alpha-crystallins

As major chaperone of eye lens, alpha-crystallin (α-Crs) is responsible for the transparency and refractive power of this region by preventing denaturation and precipitation of other proteins. As shown previously, cataract formation was positively associated with high salt intake and the elevation of blood sugar level. Here the effect of both temperature and ionic strength were studied on structure and chaperoning function of glycated and non-glycated α-Crs. While chaperone activity of these proteins was increased as function of temperature elevation, in the presence of sodium salt (0- 160 mM), it was significantly decreased. As shown by fluorescence and circular dicroism (CD) instruments, the salt induced structural alteration of α-Crs was accompanied with the exposure of hydrophobic surfaces and a transition from alpha- helical to beta-sheet structures. Moreover, the structural alterations induced by the salt were more pronounced in the case of glycated α-Crs compared to that of non-glycated protein counterpart. Overall this study shows the structural changes accompanied with lose of the chaperone activity of α-Crs induced by sodium chloride. Consequently, the obtained results may provide new evidences for the relationship between high salt intakes and cataract disease induced particularly by prolonged hyperglycemia.     

A new sesquiterpenoid from Sclerorhachis platyrachis (Boiss.) Podlech ex Rech.f

A new eudesmane-type sesquiterpenoid together with two known flavonoids were isolated from the chloroform extract of the aerial part of Sclerorhachis platyrachis. The structure of the new compound was deduced from its comprehensive spectroscopic analysis including IR, EI-MS, ¹H NMR, ¹³C NMR, DEPT, COSY, HMBC and HMQC and was shown to be 4R^*-hydroxy-6S^*-tigloyloxyeudesma-7S^*-11 (13)-en-12-oic acid (1). Finally, the structure of the new compound was unambiguously confirmed by single-crystal X-ray analysis. The structure of known compounds 2 and 3 were identified by comparison of their spectral data with those reported in the literature.