Association of Htra1 gene polymorphisms with the risk of developing AMD in Iranian population

Background:

Half of the cases of vision loss in people under 60 years of age have been attributed to age-related macular degeneration (AMD). This is a multifactorial disease with late onset. It has been demonstrated that many different genetic loci are implicated in the risk of developing AMD in different populations. In the current study, we investigated the association of high-temperature ‎requirement A-1 (HTRA1) gene polymorphisms with the risk of developing AMD in the Iranian population.

Methods:

Genomic DNA samples were extracted from 120 patients with AMD and 120 healthy age- and sex-matched controls. A 385 base-pair fragment of the HTRA1 gene promoter region was amplified using the polymerase chain reaction (PCR) technique and sequenced. The frequencies of the alleles were calculated and statistical analysis was performed using SPSS software.

Results:

Our study demonstrated that the rate of polymorphisms rs11200638 -625 G>A and rs2672598 -487T>C were significantly greater in AMD patients than in healthy controls from the Iranian population.

Conclusions:

The results of our study indicate that HTRA1 gene promoter region polymorphisms are associated with the risk of developing AMD in the Iranian population.Key Words: HTRA1, Single Nucleotide Polymorphisms, Macular Degeneration, Iran     

Studies on the Refolding Process of Recombinant Horseradish Peroxidase

Horseradish peroxidase (HRP) is an important heme-containing glyco-enzyme that has been used in many biotechnological fields. Valuable proteins like HRP can be obtained in sufficient amounts using Escherichia coli as an expression system. However, frequently, the expression of recombinant enzyme results in inclusion bodies, and the refolding yield is generally low for proteins such as plant peroxidases. In this study, a recombinant HRP was cloned and expressed in the form of inclusion bodies. Initially, the influence of few additives on HRP refolding was assessed by the one factor at a time method. Subsequently, factors with significant effects including glycerol, GSSG/DTT, and the enzyme concentration were selected for further optimization by means of the central composite design of response surface methodology (RSM). Under the obtained optimal condition, refolding increased about twofold. The refolding process was then monitored by the intrinsic fluorescence intensity under optimal conditions (0.35 mM GSSG, 0.044 mM DTT, 7 % glycerol, 1.7 M urea, and 2 mM CaCl₂ in 20 mM Tris, pH 8.5) and the reconstitution of heme to the refolded peroxidase was detected by the Soret absorbance. Additionally, samples under unfolding and refolding conditions were analyzed by Zetasizer to determine size distribution in different media.     

Antigenic Potency of LY6E in Stimulating Dendritic Cells to Elicit Tumor-Specific Responses Against Human Colorectal and Gastric Cancer Cell Lines

Early-stage gastrointestinal (GI) carcinomas are amenable malignancies, however, due to late diagnosis or lack of proper medication, alternative treatment necessitates new approaches such as dendritic cell (DC) therapy. Our previous microarray study indicated Lymphocyte antigen-6 (LY6E) as a commonly overexpressed biomarker in three lethal GI cancers, colon, gastric, and pancreatic. Therefore, we examined the antigenic potency of LY6E in stimulating DCs to elicit tumor-specific responses against human colorectal cancer (CRC) and gastric cancer (GC) cell lines HT-29 and AGS, respectively. LY6E peptide-pulsed DCs stimulated lymphocytes up to 55.9% in comparison with mature DCs (48.3%). Also, flow cytometry analysis of lymphocyte proliferation illustrated that the populations CD4⁺ and CD8⁺ were increased after treating by peptide-pulsed DCs (62.9% and 48.7% respectively). Furthermore, the cytotoxicity assay demonstrated that the 40:1 ratio of stimulated lymphocytes on AGS and HT29 cell lines was 65.1% and 66.2%, respectively. The research exposed that LY6E loaded DCs had substantial impact stimulation, proliferation, and lineage differentiation of lymphocytes. Besides, co-cultured of primed lymphocytes with AGS and HT29 cell lines exhibited cytotoxic activity. These data suggest LY6E as a potential candidate in developing DC therapy against CRC and AGS.

     

In-Silico and In-Vitro Evaluation of Antibacterial,Cytotoxic, and Apoptotic Activity and Structure of Modified CM11 Peptide

Drug-resistant infectious diseases have increased in recent years. Accordingly, plenty of researches are exploring novel approaches to overcome this problem. In this era, antimicrobial peptides have been identified as potential antibacterial agents. The Modified CM11 (mCM11) was designed with the C-terminal amidation and substitution of lysine with arginine. The designed peptide was synthesized by the solid-phase method and Rink amide p-methyl-benzhydryl amine resin. The synthesized peptide was evaluated using Mass Spectrometry (MS), High-Performance Liquid Chromatography (HPLC), and Circular Dichroism (CD). Finally, the antibacterial, cytotoxic, and apoptotic effect of the mCM11 peptide was investigated. The new peptide indicated a beta-sheet structure with a molecular weight of 1527.50 D and purity of 96%. The peptide exerted a potent antimicrobial activity against gram-negative and positive bacteria. The minimum inhibitory concentration (MIC) and minimum bacterial concentration (MBC) ranged from 16 to 64 µg/ml, and 16 to 128 µg/ml, respectively. The IC₅₀ of mCM11 was 16 µg/ml and its cytotoxicity in SH-SY5Y cell line revealed a dose-dependent manner. Also, apoptosis analysis of eukaryotic cells revealed a decline in late apoptosis and necrosis in comparison with untreated cells. The mCM11 indicated a considerable antibacterial effect against a wide range of pathogenic bacterial strains. Further, it did not represent any late apoptotic and necrosis impact on the eukaryotic cell line. All of these findings may confirm the potential role of this new peptide as an effective therapeutic agent.

     

Antimicrobial susceptibility profiles associated with bacterial meningitis among children: a referral hospital-based study in Iran

Bacterial meningitis continues to be associated with high morbidity and mortality rate worldwide, especially in the pediatric age group. This study was performed to identify the microbial etiologies of meningitis among 31 children, who were admitted in the Emergency Ward of a referral pediatric hospital in Iran. Culture identification showed that Streptococcus pneumoniae (12 subjects), Haemophilus influenzae (11 subjects) were the most common bacteria, followed by Escherichia coli (7 cases) and Neisseria meningitidis (only one case). Antibiotic susceptibility tests revealed that vancomycin had the best effect on S. pneumoniae in comparison with other antibiotics, whereas H. influenzae and E. coli were more susceptible to ceftriaxone, ceftazidime, and ceftizoxime than other antibiotics. In conclusion, despite the advances in antibiotic therapy and vaccine development, bacterial meningitis still is a health problem. S. pneumoniae, H. influenzae, and N. meningitidis are the main sources of bacterial meningitis, but other organisms such as E. coli should also be suspected, when a case is admitted to a referral pediatric hospital.     

Enhancement of antioxidant enzymes activity and expression of CAT and PAL genes in hazel (Corylus avellana L.) cells in response to low-intensity ultrasound

Induction of antioxidant systems of hazel cells by low-energy ultrasound, the potential role of hydrogen peroxide (H₂O₂) as a signaling molecule in regulation of activity of stress-related enzymes, and expression of catalase (CAT) and phenylalanine ammonialyase (PAL) genes were investigated. Suspension-cultured Corylus avellana L. cells were agitated by an ultrasonic device at 29 kHz with the power of 4 mW/cm², for 8–40 min. The activities of CAT, superoxide dismutase (SOD), and ascorbate peroxidase (APX) of treated cells increased by 4, 1.7 and 7 times of the control ones, respectively. Induction of increase in the expression of CAT gene started 24 h after the treatment with ultrasound. Significant increase also was observed in the expression of PAL gene, 6 h after exposure to ultrasound, which resulted in turn to increase of total contents of soluble phenolics, 24 h of the treatment. Exposure to ultrasound up to 20 min had no adverse effects on cell viability although it slightly increased the accumulation of H₂O₂. However, it is likely that this level of increased H₂O₂ was not deteriorative for hazel cells, but rather triggered antioxidant system and provided hazel cells a sustainable growth after ultrasound treatment.     

Chitin-Induced Gene Expression in Secondary Metabolic Pathways of Streptomyces coelicolor A3(2) Grown in Soil

Microarray analyses revealed that the expression of genes for secondary metabolism together with that of primary metabolic genes was induced by chitin in autoclaved soil cultures of Streptomyces coelicolor A3(2). The data also indicated that DasR was involved in the regulation of gene expression for chitin catabolism, secondary metabolism, and stress responses.     

Synthesis,Characterization, Molecular Docking,and Biological Activities of Some Natural and Synthetic Urolithin Analogs

Urolithins (that is, hydroxy substituted benzo[c]chromen‐6‐one derivatives) are formed within the gastrointestinal tract following to the exposure to various ellagitannin rich diet, particularly involving pomegranate, nuts, and berries. Regarding the bioavailability deficiency of ellagitannins, the biological activities obtained through the extracts of these dietaries are attributed to the urolithin compounds, since they are bioavailable. Particularly, there are studies indicating the importance of ellagitannin‐rich food for protective and alternative treatment of Alzheimer's Disease (AD). From this perspective, within this study, the major urolithins (that is, urolithins A and B), their methyl ether metabolites, as well as some synthetic urolithin analogs have been synthesized and screened for their biological activities in various enzyme inhibition (acetylcholinesterase, butyrylcholinesterase, monoamine oxidase B, cyclooxygenase 1, and cyclooxygenase 2) and antioxidant (DPPH radical scavenging) assay systems. The results pointed out the potential of urolithins to act as inhibitors on these receptors. Docking studies were also performed to investigate the possible interactions.     

Resource recovery from low strength wastewater in a bioelectrochemical desalination process

In this research, low strength synthetic wastewaters with chemical oxygen demand less than 300 mg L^?1 were treated at different concentrations in a bioelectrochemical desalination process. A process optimization model was utilized to study the performance of the photosynthetic bioelectrochemical desalination process. The variables include substrate (chemical oxygen demand) concentration, total dissolved solids, and microalgae biomass concentration in the cathode chamber. Relationships between the chemical oxygen demand concentration, microalgae, and salt concentrations were evaluated. Power densities and potential energy benefits from microalgal biomass growth were discussed. The results from this study demonstrated the reliability and reproducibility of the photosynthetic microbial desalination process performance followed by a response surface methodology optimization. This study also confirms the suitability of bioelectrochemical desalination process for treating low substrate wastewaters such as agricultural wastewaters, anaerobic digester effluents, and septic tank effluents for net energy production and water desalination.     

Phasic Contractions in Urinary Bladder from Juvenile versus Adult Pigs

Aims

Alterations in properties of the bladder with maturation are relevant physiologically and pathophysiologically. The aim of this study was to investigate alterations in bladder properties with maturation in juvenile vs. adult pig, focussing on differences between layers of the bladder wall (mucosa vs. detrusor) and the presence and functional contribution of interstitial cells (ICs).

Methods

Basal and cholinergic-induced phasic contractions (PCs) in mucosal and denuded-detrusor strips from juvenile and adult pigs were assessed. Expression of c-kit, a marker of ICs, was investigated in the mucosa and the detrusor layers of the pig bladder. The functional role of ICs in mediating PCs was examined using imatinib.

Results

Mucosal strips from juvenile and adult pig bladders demonstrated basal PCs whilst denuded-detrusor strips did not. PCs of mucosal strips from juvenile pigs were significantly greater than those from adult bladders. Immunoreactivity for c-kit was detected in mucosa and detrusor layers of pig bladder. Histological studies demonstrated a distinct layer of smooth muscle between the urothelium and bladder detrusor, termed the muscularis mucosa. Imatinib was only effective in inhibiting PCs in mucosal strips from juvenile pigs. Imatinib inhibited the carbachol-induced PCs of both juvenile and adult denuded-detrusor strips, although strips from juvenile bladders demonstrated a trend towards being more sensitive to this inhibition.

Conclusions

We confirm the presence of c-kit positive ICs in pig urinary bladder. The enhanced PCs of mucosal strips from juvenile animals could be due to altered properties of ICs or the muscularis mucosa in the bladders of these animals.