Characterization and biocompatibility evaluation of acellular rat skin scaffolds for skin tissue engineering applications.

Utilization of acellular scaffolds, extracellular matrix (ECM) without cell content, is growing in tissue engineering, due to their high biocompatibility, bioactivity ad mechanical support. Hence, the purpose of this research was to study the characteristics and biocompatibility of decellularized rat skin scaffolds using the osmotic shock method. First, the skin of male Wistar rats was harvested and cut into 1 × 1 cm2 pieces. Then, some of the harvested parts were subjected to the decellularization process by applying osmotic shock. Comparison of control and scaffold samples was conducted in order to assure cell elimination and ECM conservation by means of histological evaluations, quantification of biochemical factors, measurement of DNA amount, and photographing the ultrastructure of the samples by scanning electron microscopy (SEM). In order to evaluate stem cell viability and adhesion to the scaffold, adipose-derived mesenchymal stem cells (AD-MSCs) were seeded on the acellular scaffolds. Subsequently, MTT test and SEM imaging of the scaffolds containing cultured cells were applied. The findings indicated that in the decellularized scaffolds prepared by osmotic shock method, not only the cell content was removed, but also the ECM components and its ultrastructure were preserved. Also, the 99% viability and adhesion of AD-MSCs cultured on the scaffolds indicate the biocompatibility of the decellularized skin scaffold. In conclusion, decellularized rat skin scaffolds are biocompatible and appropriate scaffolds for future investigations of tissue engineering applications.

Risk of Gastric Cancer is Highly Dependent on Type of First-Degree Family Member Affected by Cancer: Lessons from a High-Risk Population in Iran.

Background: Family history of gastric cancer (GC) in first-degree relatives may increase the risk of GC. This study aimed to assess how family history of GC in first-degree relatives really affects the risk of GC in an extremely high-risk population. Methods: A large population-based case-control study was carried out on 1222 incident GC cases and 1235 controls in Ardabil Province-a high-risk area in North-West Iran-to assess the associations of GC family history in first-degree relatives with the risk of GC (2003-2017). Results: GC family history did not significantly associate with the risk of GC overall (ORadj=1.09, 95% CI: 0.80-1.47, P=0.589). It found no significant association of GC family history in a parent, and in a father, mother, and sister separately, with the risk of GC. However, GC risk was significantly associated with a history of GC in a sibling (ORadj=1.61, 95% CI: 1.11-2.35, P=0.013), especially brother (ORadj=2.24, 95% CI: 1.41-3.64, P=0.0008). The risk was greatly increased in subjects with two or more affected brothers (ORadj =5.56, 95% CI: 2.33-14.20, P=0.0002). Conclusion: We did not find a familial tendency to cardia GC and non-cardia GC as well as histopathologic features. Determining the type of first-degree relationships with GC may, therefore, be more important than assessing family history alone for predicting the risk of GC in this high-risk area.

Effect of Azithromycin on Sciatic Nerve Injury in the Wistar Rats.

After a severe peripheral nerve injury, complete functional recovery is rare. Modulating the inflammatory response could be an effective way to enhance peripheral nerve regeneration. The present study aimed to determine the effect of azithromycin on functional recovery following sciatic nerve crush in Wistar rats. 40 male Wistar rats were used in four groups, including: the negative control, sham, and two groups of azithromycin (15 and 150 mg/kg/day) (n = 10).The rats' right sciatic nerve was crushed using a non-serrated clamp. In experimental groups, animals were treated with azithromycin (15 and 150 mg/kg/day) for 7 days. Then, sensory-motor functions were evaluated over eight weeks. Real-time PCR was used to measure the expression of NGF and BDNF genes. At the end of the 4th week, the sensory recovery accelerated in the azithromycin-treated rats so that the reaction times in the groups treated with 15 mg/kg and 150 mg/kg doses of azithromycin reached 5.14 s and 6.61 s, respectively, which were significantly lower than the 12 s in the negative control group (P < 0.05).Eventually, the mean SFI values in the negative control and both azithromycin-treated groups recovered to preoperative levels in the 8th week, with no significant difference between the sciatic lesion groups. Findings showed a seven-day course of azithromycin administered immediately after a sciatic nerve crush could accelerate regeneration and improve motor and sensory function recovery compared to negative controls. These significant effects were observed in both the azithromycin 15 mg/kg and the azithromycin 150 mg/kg treatment groups. Azithromycin treatment upregulated the expression of NGF and BDNF genes in crushed sciatic nerve. Our findings suggest that a seven-day treatment of azithromycin after a sciatic nerve injury could accelerate the regeneration process and improve functional recovery.

Glibenclamide promoted functional recovery following sciatic nerve injury in male Wistar rats.

The impact of peripheral nerve damage on a patient's quality of life is severe. The most frequent peripheral nerve crush damage is a sciatic nerve injury. Previous research has shown that glibenclamide (GB) has neuroprotective properties in a variety of oxidative stress-related disorders, including Alzheimer and Parkinson. The goal of this study was to see how GB affected nerve regeneration and improved function of the sciatic nerve in a rat model following a crush injury. We evaluated motor function, sensory recovery, gene expression, and histomorphometry following damage at different time points. Additionally, we assessed atrophy in the gastrocnemius muscle using histology and mass ratio analyses. Our results suggest that 2, 4, 6, and 8 weeks following glibenclamide therapy, promotes the recovery of motor and sensory function in the injured site. Following glibenclamid injection, the mRNA levels of neurotrophic factors (NGF and BDNF) are raised. According to histomorphometry assessment, glibenclamide injection also increased the number of myelinated fibers while decreasing their thickness. These results showed that glibenclamide therapy by decreasing the proinflammatory and oxidant factors may enhance the nerve regeneration. It is clear that more research is needed to confirm these findings.

Estradiol modulated differentiation and dynamic growth of CD90+ spermatogonial stem cells toward Sertoli-like cells.

Mouse CD90+ SSCs were enriched using the MACS technique and incubated with different doses of estradiol, ranging from 0.01 ng/mL to 500 µg/mL, for 7 days. The viability of SSCs was determined using an MTT assay. The combined effects of estradiol plus Sertoli cell differentiation medium on the orientation of SSCs toward Sertoli-like cells were also assessed. Using immunofluorescence imaging, we monitored protein levels of Oct3/4 after being exposed to estradiol. In addition, protein levels of testosterone, TF, and ABP were measured using ELISA. The expression of Sertoli cell-specific genes such as SOX9, GATA4, FSHR, TF, and ESR-1 and -2 was monitored using real-time PCR assay, and the effects of 14-day injection of estradiol on sperm parameters and Oct3/4 positive progenitor cells in a model of mouse were determined. Data showed that estradiol increased the viability of mouse SSCs in a dose-dependent manner compared to the control (p < 0.05). Along with these changes, cells displayed morphological changes and reduced Oct3/4 transcription factor levels compared to the control SSCs. 7-day incubation of SSCs with estradiol led to the up-regulation of SOX9, GATA4, FSHR, TF, and ESR-1 and -2, and levels of testosterone, TF, and ABP were increased compared to the control group (p < 0.05). The in-vivo examination noted that estradiol reduced sperm parameters coincided with morphological abnormalities (p < 0.05). Histological examination revealed pathological changes in seminiferous tubules and reduction of testicular Oct3/4+ progenitor cells. In conclusion, estradiol treatment probably can induce Sertoli cell differentiation of SSCs while exogenous administration leads to testicular progenitor cell depletion and infertility in long term.

Contrasting association of Helicobacter pylori oipA genotype with risk of peptic ulceration and gastric cancer.

Helicobacter pylori OipA (outer inflammatory protein A) is an outer membrane protein that involves in the binding and colonization of the bacterium in the stomach. The oipA status is associated with the risk of peptic ulcerations (PUs) and gastric cancer (GC) diseases. However, the association trend with PUs compared to GC is often different and highly challenging. We therefore aimed to determine the presence of this genotype in Iranian strains and assess its association with the risk of PUs and GC in a larger number of samples. A total of 319 strains were obtained from 172 patients with non-atrophic gastritis (NAG), 52 with PUs and 95 with GC. The prevalence of the oipA+vs. oipA- genotype was 67.7% (216/319). The total frequency of the oipA+vs. oipA- genotypes in NAG, PUs, GC, non-peptic ulceration (including NAG and GC), and non-tumor (including NAG and PUs) groups was 121/172 (70.3%), 50/52 (96.2%), 45/95 (47.4%), 166/267 (62.2%), and 171/224 (76.3%), respectively. In multiple logistic regression analysis, the oipA+vs. oipA- genotype showed a strong direct association with PUs; the ORadj (95% CI) was 18.751 (4.421-79.531), (p = 0.00007). In contrast, it had a significant reverse association with GC; the ORadj (95% CI) was 0.330 (0.179-0.607), (p = 0.00036). In the present study, we interestingly found a contrasting association of the H. pylori oipA genotype with the risk of PUs and GC in Iran. Therefore, the contrasting effect of this genotype may emphasize its independent role in predicting clinical outcomes.

SNP-SNP interactions of oncogenic long non-coding RNAs HOTAIR and HOTTIP on gastric cancer susceptibility.

Genetic variants within oncogenic long non-coding RNAs HOTAIR and HOTTIP may affect their gene expression levels, thereby modifying genetic susceptibility to gastric cancer (GC). In a hospital-based study in Ardabil-a very high-risk area in North-West Iran, 600 blood samples from 300 GC patients and 300 healthy controls were recruited for genotyping. Seven HOTAIR (i.e., rs17720428, rs7958904, rs1899663, and rs4759314) and HOTTIP (i.e., rs3807598, rs17501292, and rs1859168) 'tag' single nucleotide polymorphisms (SNPs) were genotyped by the Infinium HTS platform. The rs17720428, rs7958904, and rs1899663 tagSNPs significantly increased GC risk under dominant models by 1.5-, 1.57-, and 1.5-fold, respectively. The G-C-T-A haplotype of HOTAIR tagSNPs increased the risk of GC by 1.31-fold. No significant association was found between HOTTIP SNPs and the risk of GC. HOTAIR and HOTTIP variants were also not associated with any clinicopathologic characteristics. The SNP-SNP interaction of HOTAIR rs17720428/rs7958904 with HOTTIP rs1859168 was associated with an increased risk of GC (rs17720428 TG-rs1859168 CC, OR = 1.76; rs7958904 GC-rs1859168 CC, OR = 1.85; rs7958904 CC-rs1859168 CC, OR = 1.86). Interestingly, the SNP-SNP interaction of HOTAIR rs1899663 with HOTTIP rs1859168 strongly increased the risk of GC (rs1899663 GT-rs1859168 CC, OR = 4.3; rs1899663 TT-rs1859168 CC, OR = 9.37; rs1899663 TT-rs1859168 CA, OR = 6.59). We showed that the HOTAIR rs17720428, rs7958904, and rs1899663 tagSNPs and their interactions with the HOTTIP rs1859168 polymorphism significantly increased the risk of GC. Specifically, novel SNP-SNP interactions between HOTAIR and HOTTIP tagSNPs have a larger impact than individual SNP effects on GC risk, thereby providing us with valuable information to reveal potential biological mechanisms for developing GC.

Rosuvastatin enhanced functional recovery after sciatic nerve injury in the rat.

Posttraumatic nerve recovery remains a challenge in regenerative medicine. As such, there is a need for agents that limit nerve damage and enhance nerve regeneration. Here we investigate rosuvastatin, a 3-hydroxy-3-methylglutaryl coenzyme (HMG-CoA) reductase inhibitor, with anti-inflammatory and antioxidant properties. We explore its neuroprotective properties on sciatic nerve crush injury in male Wistar Rats. Rats were subjected to crush injury to the left sciatic nerve using a vessel clamp for 30 s. Rosuvastatin or vehicle was prepared daily and administrated by oral gavage for seven days post-injury. In rosuvastatin treatment groups, rosuvastatin was administrated at the doses of (5 or 10 mg/kg) in the treatment group. The control group was given a vehicle in the same manner. Behavioral, electrophysiological, morphological and molecular parameters were examined during the recovery process. Chronic administration of rosuvastatin at all doses after sciatic nerve crush markedly promoted nerve regeneration and significantly accelerated motor function recovery (P < 0.05). Electrophysiological, morphological and molecular parameters also improved in the rosuvastatin treatment groups compared to the controls. These findings suggest that neuroprotective effects of rosuvastatin could be due to its antioxidant and anti-inflammatory activity. It is clear that more research is needed to confirm these findings.

Biosynthesis, characterization and evaluation of the supportive properties and biocompatibility of DBM nanoparticles on a tissue-engineered nerve conduit from decellularized sciatic nerve.

In this study, we examined the supporting effects of nano-demineralized bone matrix on the cultivation of Wharton's jelly stem cells on acellularized nerve scaffold. Demineralized bone matrix nanoparticles were prepared and characterized by several experiments. Decellularized sciatic nerve scaffolds were prepared and their efficiency was evaluated using histological stainings and biomechanical testing. Results of histological staining indicated that the integrity of the extra cellular matrix components was preserved. Also, the growth and viability of WJSCs on the scaffolds were significantly higher in DBM nanoparticle groups. We conclude that supportive properties of nano-DBM groups showed better cell viability and a suitable microenvironment for proliferation, retention, and adhesion of cells compared with other groups.

Targeting signaling pathways of VEGFR1 and VEGFR2 as a potential target in the treatment of breast cancer.

Tumor angiogenesis allows tumor cells to grow and migrate toward the bloodstream and initiate metastasis. The interactions of vascular endothelial growth factors (VEGF) A and B, as the important regulating factors for blood vessel growth, with VEGFR1 and VEGFR2 trigger angiogenesis process. Thus, preventing these interactions led to the effective blockade of VEGF/VEGFRs signaling pathways. In this study, the inhibitory effect of a 23-mer linear peptide (VGB4), which binds to both VEGFR1 and VEGFR2, on VEGF-stimulated Human Umbilical Vein Endothelial Cells (HUVECs) and highly metastatic human breast cancer cell MDA-MB-231 proliferation was examined using MTT assay. To assess the anti-migratory potential of VGB4, HUVECs and also MDA-MB-231 cells wound healing assay was carried out at 48 and 72 h. In addition, downstream signaling pathways of VEGF associated with cell migration and invasion were investigated by quantification of mRNA and protein expression using real-time quantitative PCR and western blot in 4T1 tumor tissues and MDA-MB-231 cells. The results revealed that VGB4 significantly impeded proliferation of HUVECs and MDA-MB-231 cells, in a dose- and time-dependent manner, and migration of HUVECs and MDA-MB-231 cells for a prolonged time. We also observed statistically significant reduction of the transcripts and protein levels of focal adhesion kinase (FAK), Paxillin, matrix metalloproteinase-2 (MMP-2), RAS-related C3 botulinum substrate 1 (Rac1), P21-activated kinase-2 (PAK-2) and Cofilin-1 in VGB4-treated 4T1 tumor tissues compared to controls. The protein levels of phospho-VEGFR1, phospho-VEGFR2, Vimentin, ß-catenin and Snail were markedly decreased in both VGB4-treated MDA-MB-231 cells and VGB4-treated 4T1 tumor tissues compared to controls as evidenced by western blotting. These results, in addition to our previous studies, confirm that dual blockage of VEGFR1 and VEGFR2, due to the inactivation of diverse signaling mediators, effectively suppresses tumor growth and metastasis.

Prevalence of K-RAS mutations and CA125 tumor marker in patients with ovarian carcinoma.

BACKGROUND: Ovarian carcinoma is one of the leading causes of cancer-related death among females. K-ras codon 12 mutations are commonly occurring mutations in different types of cancers and leads to resistance against anti-EGFR therapeutics. Hence, determination of mutations in k-ras gene is crucial for predicting response to anti-EGFR therapies. This study aimed to evaluate the prevalence of k-ras gene mutations and CA125 tumor marker in patients with ovarian carcinoma in Tabriz city. METHODS: Genomic DNA was extracted from 67 tissues pertained to women with ovarian carcinoma. Mutations in codon 12 and 13 of k-ras gene were analyzed by Nested PCR and RFLP methods. Titer of CA125 tumor marker was determined by ELISA. RESULTS: Among the 67 patients, 7 patients (10.4%) had mutation in k-ras codon 12 while none of them had mutation in k-ras codon 13. Based on the results, the frequency of various genotypes were 89.55%, 10.44%, and 0%, for GG, GA, and AA alleles, respectively. The p-value for stage I and Grade I tumors were 0.022 and 0.04, respectively, indicating a statistically significant correlation between codon 12 mutation and stage I and Grade I tumors. Furthermore, we found significant correlations among CA125 tumor marker titers and histological grade (p<0.000) and stage (p<0.000). The mean serum CA125 tumor marker levels in malignant carcinomas were 499.84 U/ml. CONCLUSION: The results in this study indicated high prevalence of k-ras codon 12 mutations and high titer of CA125 tumor marker in patients with ovarian carcinoma in the study region.

Hair follicle stem cells differentiation into bone cells on collagen scaffold.

The hair follicle is a dynamic structure which contains different niches for stem cells, therefore; it has been considered as valuable and rich sources of stem cells, due to easy access, multipotency and non-oncogenic properties. In the present study, the differentiation capacities of hair follicle stem cells into bone cells on the natural collagen scaffolds were investigated. The stem cells were extracted from the hair follicle bulge area of male Wistar rats' whisker and cultured until 3rd passage, then osteogenic differentiations were induced by culturing the cells in the specific osteogenic medium. After 3 weeks, the differentiation parameters, including morphological changes, levels of calcification and expression of the bone specific genes were detected. The hydrogel preparation and scaffold fabrication was carried out using the extracted collagen and was studied by scanning electron microscope. Comparison of the stem cells' growth and changes on the scaffold and non-scaffold conditions showed that, in the both situation, the cells revealed differentiation signs of osteocytes, including large and cubic morphology with a star-shaped nucleus. Staining by Alizarin-red and Von-Kossa methods showed the presence of red and black calcium mass on the scaffold. Expression of the osteopontin and alkaline phosphatase genes confirmed the differentiation. Considerable porosity in the surface of the scaffold was recorded by scanning electron microscopy, which made it convenient for cells' attachment and growth. The data showed that the bulge stem cells possess significant capacity for osteoblastic differentiation and collagen scaffolds were found to be an appropriate matrix for growth and differentiation of the cell.

Unique constellations of five polymorphic sites of Helicobacter pylori vacA and cagA status associated with risk of gastric cancer.

Helicobacter pylori possesses virulence genes that are involved in the pathogenesis of the bacterium. There are little data regarding all constellations of five polymorphic sites of H. pylori vacA and cagA status. We therefore aimed to i) find any associations between H. pylori vacA alleles (s1/s2, m1/m2, i1/i2, d1/d2, and c1/c2) and cagA status and ii) determine the frequency of all five-genotype combinations of the vacA alleles with and without cagA gene, and their associations with risk of gastric cancer (GC). A total of 290 Iranian H. pylori isolates from gastrointestinal patients were obtained successfully by the cultivation of biopsies and genotyped. The patients included 144/290 with non-atrophic gastritis (NAG), 57/290 with peptic ulcer disease (PUD), and 89/290 with GC. We found that each of the vacA m1-, i1-, d1-, and c1-genotypes was significantly associated with cagA+ status. The odds ratio(OR) and 95% confidence interval (95% CI) was 2.316 (1.241-4.301) for cagA+/vacA m1, 2.764 (1.540-4.960) for cagA+/vacA i1, 4.288 (2.305-7.977) for cagA+/vacA d1, and 2.639 (1.488-4.680) for cagA+/vacA c1, respectively. In this study, 43 five- and six-genotype combinations were found among 224 strains. The highest frequencies were observed for vacA s1m2i2d2c2 (85/224, 37.9%), s1m2i2d2c2/cagA (48/222, 21.6%), s1m1i1d1c1 (40/224, 17.9%) and s1m1i1d1c1/cagA (35/222, 15.8%). Logistic regression analysis showed that vacA s1m1i1d1c1, s1m2i1d2c1, s1m2i2d2c1, and s1m2i2d2c1/cagA had a high prevalence in GC patients compared to non-atrophic gastritis patients (p < .05). The ORs and 95% CI were 2.433 (1.070-5.531), 11.524 (1.253-106.023), 4.200 (1.261-13.993), and 6.263 (1.494-26.256), respectively. These results were also confirmed when the controls were non-tumors (NAG/PUD). We found novel five- and six-genotype combinations associated with the risk of GC. These associations seem to be strongly dependent on the presence of c1-type of vacA. Therefore, analysis of all combined genotypes of the vacA alleles and cagA status may play a significant role in determining H. pylori-related clinical outcomes.

Risk factors predisposing to cardia gastric adenocarcinoma: Insights and new perspectives.

Recent decades have seen an alarming increase in the incidence of cardia gastric adenocarcinoma (CGA) while noncardia gastric adenocarcinoma (NCGA) has decreased. In 2012, 260 000 CGA cases (age-standardised rate (ASR); 3.3/100 000) and 691 000 NCGA cases (ASR; 8.8/100 000) were reported worldwide. Compared with women, men had greater rates for both the subsites, especially for CGA. Recently, four molecular subtypes of GC have been proposed by the Cancer Genome Atlas (TCGA) and the Asian Cancer Research Group (ACRG); however, these classifications do not take into account predisposing germline variants and their possible interaction with somatic alterations in carcinogenesis. The etiology of adenocarcinoma of the cardia and the gastroesophageal junction (GEJ) is not known. It is thought that CGA is distinct from adenocarcinomas located in the esophagus or distal stomach, both epidemiologically and biologically. Moreover, CGA is often identified in the advanced stage having a poor prognosis. Therefore, understanding the risk and the role of predisposing factors in etiology of CGA can inform clinical practice and counseling for risk reduction. In this paper, we showed that GC family history, lifestyle, demographics, gastroesophageal reflux disease, Helicobacter pylori infection, and multiple genetic and epigenetic risk factors as well as several predisposing conditions may underlie susceptibility to CGA. However, several genome-wide association studies (GWASs) should be conducted to identify novel high-penetrance genes and pathways as well as causal germline variants predisposing to CGA. They must include different ethnic groups, especially from high-incidence countries for CGA, because some risk loci are ancestry-specific. In parallel, statistical methods can be developed to identify cancer predisposition genes (CPGs) from tumor sequencing data. It is also necessary to find novel long noncoding RNAs related to the risk of CGA. Taken altogether, new cancer risk prediction models, including all genetic and nongenetic factors influencing risk, should be developed to facilitate risk assessment, disease prevention, and early diagnosis and intervention of CGA in the future.

Inverse association of Helicobacter pylori cagPAI genotypes with risk of cardia and non-cardia gastric adenocarcinoma.

Iran is a high-risk country for cardia gastric adenocarcinoma (CGA) in Central Asia, with an incidence rate five times the average global rate, and shows a high infection rate for Helicobacter pylori (69%). The aim was to examine the associations of multiple H. pylori cagPAI genotypes (ie cagH, cagL, cagG, and orf17) with the risk of CGA, non-CGA, and different histological types of GA in Iran. A large number of H. pylori strains (N = 336) were successfully cultured and genotyped. Histopathological evaluations were performed. The analysis showed an inverse association between the cagH+ genotype and the risk of CGA and intestinal-type gastric adenocarcinoma (IGA) (adjusted ORs; 0.312 and 0.283, respectively), where the controls were nontumors. The orf17+ genotype decreased the risk of non-CGA and diffuse-type gastric adenocarcinoma (DGA)(adjusted ORs; 0.310 and 0.356, respectively). When the controls were those with nonatrophic gastritis, the cagG+ genotype was negatively associated with the risk of CGA, non-CGA, IGA, and DGA (adjusted ORs; 0.324, 0.366, 0.306, and 0.303, respectively). We did not find such a significant association for the cagL+ genotype in multiple logistic regression analysis. Combination of the vacA c2 and cagPAI genotypes further decreased the risk estimates for GAs. This study showed the reverse association of H. pylori cagPAI genotypes-cagH+ and cagG+ -with the risk of CGA in male patients aged ≥ 55 in Iran. Presence of the vacA c2 genotype in combination with cagPAI genotypes showed strong inverse associations with the risk of CGA and non-CGA. These findings may reveal a coordinated relationship between the vacA c2 and cagPAI genotypes.

A peptide mimicking the binding sites of VEGF-A and VEGF-B inhibits VEGFR-1/-2 driven angiogenesis, tumor growth and metastasis.

Interfering with interactions of vascular endothelial growth factors (VEGFs) with their receptors (VEGFRs) effectively inhibits angiogenesis and tumor growth. We designed an antagonist peptide of VEGF-A and VEGF-B reproducing two discontinuous receptor binding regions of VEGF-B (loop 1 and loop3) covalently linked together by a receptor binding region of VEGF-A (loop3). The designed peptide (referred to as VGB4) was able to bind to both VEGFR1 and VEGFR2 on the Human Umbilical Vein Endothelial Cells (HUVECs) surface and inhibited VEGF-A driven proliferation, migration and tube formation in HUVECs through suppression of ERK1/2 and AKT phosphorylation. The whole-animal fluorescence imaging demonstrated that fluorescein isothiocyanate (FITC)-VGB4 accumulated in the mammary carcinoma tumors (MCTs). Administration of VGB4 led to the regression of 4T1 murine MCT growth through decreased expression of p-VEGFR1 and p-VEGFR2 and abrogation of ERK1/2 and AKT activation followed by considerable decrease of tumor cell proliferation (Ki67 expression) and angiogenesis (CD31 and CD34 expression), induction of apoptosis (increased p53 expression, TUNEL staining and decreased Bcl2 expression), and suppression of metastasis  (increased E-cadherin and decreased N-cadherin, NF-κB and MMP-9 expression). These findings indicate that VGB4 may be applicable for antiangiogenic and antitumor therapy.

Enhancing the apoptotic effect of IL-24/mda-7 on the human hepatic stellate cell through RGD peptide modification.

Induction of apoptosis or quiescent hepatic stellate cells (HSCs) can be an attractive molecular strategy due to the importance of activation of HSCs during hepatic fibrogenesis. Interleukin-24/melanoma differentiation-associated gene-7 (IL-24/mda-7) is a cytokine that has attracted a great deal of attention in the tumor killing as well as pathophysiology of the diseases. In this study, the Pro-apoptotic and senescence inductive properties of IL-24/mda-7 were assessed in human-derived HSCs. Three plasmids expressing natural mda-7, peptide modified version, mda-7-RGD genes beside a recombinant IL-24 protein, were added or transfected into activated LX-2 cells. Cell viability and the amount of apoptosis were analyzed using MTT and Annexin V staining method, respectively. Hence, the expression levels of apoptotic genes and PPARγ in different groups were also compared by real-time PCR analysis. Furthermore, the senescence effect of IL-24/mda-7 by a ß-galactosidase (SA-ß-gal) senescence assay, was evaluated. The viability assessment showed that pmda-7-RGD had the most significant growth inhibitory effect when compared to the control group, pcDNA3.1 (P = 0.0002). The apoptosis analysis also revealed a significant impact of different mda-7 forms in apoptosis induction. The measuring of cell senescence also indicated that IL-24/mda-7 in plasmid and protein forms exhibited a senescence inductive activity as determined by an increase in PPARγ gene expression and beta-galctosidase activity. In conclusion, our findings demonstrated that both endogenous and soluble forms of IL-24/mda-7 induced apoptosis and senescence in activated LX-2 cells and more importantly, fusion of RGD peptide to this cytokine enhanced these activities. So, RGD-modified IL-24/mda-7 could be a suitable candidate for further molecular therapy of fibrosis.

Are Helicobacter pylori highly cytotoxic genotypes and cardia gastric adenocarcinoma linked? Lessons from Iran.

BACKGROUND: Although the most extensive studies revealed the role of H. pylori VacA and CagA toxins in the development of gastric adenocarcinoma, the magnitude of this association and the correlations of vacA mosaicism and cagA status with cardia gastric adenocarcinoma (CGA) still remain controversial. OBJECTIVE: We aimed to examine the linkage of H. pylori highly cytotoxic genotypes to CGA in Iranian populations as a model. METHODS: A total of 601 Iranian patients were enrolled. Biopsies were cultured, genotyped, and anatomically and histologically classified. RESULTS: The vacA c1 genotype, but not cagA status, showed a strong association with the risk of both CGA and non-cardia adenocarcinoma (NCGA), whether the controls were non-tumors, as those with either non-atrophic gastritis or peptic ulcerations, (the OR (95%CI) was 14.11 (4.91-40.52) and 9.59 (4.06-22.65), respectively) or those with NAG (the OR (95%CI) was 10.71 (3.49-32.82) and 8.11 (3.26-20.16), respectively). The vacA c1/cagA+ genotype was significantly associated with an increased risk of NCGA, whether the controls were non-tumors or those with NAG; the adjusted risk was 4.706 (1.41-15.67) and 4.85 (1.42-16.51), respectively. CONCLUSIONS: The H. pylori vacA c1 genotype, but not cagA status, might be the first important bacterial biomarker for predicting the cardia adenocarcinoma risk in male patients aged ⩾ 55 in Iran.

A study on the modulation of alpha-synuclein fibrillation by Scutellaria pinnatifida extracts and its neuroprotective properties.

Aggregation of alpha-synuclein (α-SN) is a key pathogenic event in Parkinson's disease (PD) leading to dopaminergic degeneration. The identification of natural compounds inhibiting α-SN aggregation may have a major role in treating PD. Different Scutellaria species are known as valuable medicinal plants, primarily due to their high flavonoid levels. Scutellaria pinnatifida (S. pinnatifida) is endemic to Iran; however, the knowledge of its pharmaceutical properties is limited. Here we report that S. pinnatifida extracts have an anti-fibrillation effect on α-SN aggregation and neuroprotective properties on PC12 and primary dopaminergic neurons. Treatment during α-SN fibril formation with S. pinnatifida extracts showed that the extractions performed with dichloromethane (DCMEx) and n-butanol (BuOHEx) strongly inhibited α-SN fibrillation. TLC-based analysis revealed that S. pinnatifida contains a great amount of flavonoids with high antioxidant properties as shown using a radical scavenging assay. Further analysis using HPLC and Mass spectroscopy on the DCMEx revealed the presence of baicalein in this extract. We then selected the more efficient extracts based on cell viability and ROS scavenging on PC12 cells and tested their neuroprotective properties on primary dopaminergic neurons. Our results showed the extracts strongly protected against α-SN oligomers. Surprisingly, they also neutralized the severe toxicity of paraquat. Therefore, S. pinnatifida may be a potential valuable medicinal herb for further studies related to the treatment of PD.

The pattern of IL-24/mda-7 and its cognate receptors expression following activation of human hepatic stellate cells.

Activation of hepatic stellate cells (HSCs) is the pivotal event during liver fibrosis. Interleukin (IL)-24/melanoma differentiation-associated gene-7 (mda-7) has attracted attention in the pathophysiology of some diseases, while its role in activation/suppression of human HSCs is still unclear. It is important to elucidate whether the expression levels of the IL-24/mda-7 protein and its receptors in HSC cells are changed following activation. LX-2 cells, a human hepatic stellate cell line were activated by a combination of leptin and serum starvation. The activation state was evaluated through measuring the mRNA expression of profibrotic molecules, collagen-I, TIMP metalloproteinase inhibitor-1 and transforming growth factor-ß. The expression of IL-24/mda-7 was assessed in mRNA and protein levels by reverse transcription-quantitative polymerase chain reaction (RT-qPCR) and ELISA methods, respectively. Hence, the amount of IL-22R1 and IL-20R2 subunit expression was also compared in activated and normal LX-2 cells by RT-qPCR. The expression level of IL-24/mda-7 and its cognate receptors was detectable both in the normal and activated LX-2 cell line. Furthermore, in activated LX-2, a significant increase of IL24 expression either on IL-22R1 and IL-20R2 subunits was also noticeable in comparison to normal cells. The activation state of LX-2 cells caused significant changes of IL-24/mda-7 and its receptors expression. In addition, the elevation in IL-24/mda-7 during LX-2 cell activation, suggested that IL-24/mda-7 and its cognate receptors serve a possible role in the development of the fibrosis process. Therefore, IL-24/mda-7 and relevant signaling pathways may be employed as a target for fibrosis treatment.