Search

Mesenchymal stem cells (MSC) from bone marrow hold great potential as a cell source for cartilage repair. The objective of our study was differentiation of MSC toward chondrocyte by using Nanofiber-based polyethersulfone (PES) scaffold and also enhanced chondrogenic differentiation of BMSC in vitro. MSCs were harvested from bone marrow of human and PES scaffold was fabricated via Electrospinning. The isolated cells were cultured on the PES scaffold and scaffold free method. After 21 days, Real-time PCR was performed to evaluate the cartilage-specific genes in the mRNA levels. Also, in order to confirm our results, we have done immunocytochemistry and SEM imaging. Flowcytometry confirmed the nature of the isolated adherent cells. Immunocytochemistry and SEM imaging confirmed the differentiation of MSC toward chondrocyte. Also, real time PCR showed a significant increased gene expression of collagen type II and aggrecan on the PES scaffold method when compared to the mRNA levels measured in scaffold free method. Down regulation of Collagen type I was observed in PES scaffold compared to scaffold free at day 21. Also, both methods showed a similar pattern of expression of SOX9. Our results showed that PES scaffold maintains BMSC proliferation and differentiation, and can significantly enhance chondrogenic differentiation of BMSC. PES scaffold seeded BMSC showed the highest capacity for differentiation into chondrocyte-like cells. [ABSTRACT FROM AUTHOR]

CD24 is a heavy glycosylated protein that is overexpressed in numerous cancer and cancer stem cells. It is involved in the development, invasion, and metastasis of the cancer cells. CD24 can be considered in targeted cancer therapy as a new target. Here, phage display technology was used for the identification of novel scFv antibodies against CD24. To do so, the CD24 protein was expressed and purified from a stable transgenic CHO cell line. The cells were developed by using a CD24 encoding construct, which targets the 18S rRNA gene. The recombinant CD24 was used in the biopanning process. Four rounds of biopanning were performed with the Tomlinson J library. The polyclonal phage ELISA and DNA sequencing of the selected colonies confirmed that specific binders have been enriched during the biopanning process. Based on the DNA sequencing results, three phage-displaying scFv were obtained and their specificity to CD24 was verified in the monoclonal phage ELISA. The results showed that clone 3 (called J d3 ) has the highest affinity to CD24 and also is the most abundant clone (80%) among the three clones. Therefore, J d3 was selected for further analyses. In order to produce soluble J d3 -scFv, the phage was transfected to E. coli BL21 pLysS . Soluble J d3 scFv was expressed and purified successfully. The soluble J d3 scFv showed appropriate affinity to recombinant CD24 in the ELISA analysis. Also, the immunocytochemistry experiment showed that the purified J d3 scFv can bind to the CD24 expressing A549 cells. So, the scFv may be useful in the targeted delivery of drugs or diagnostic nanoparticles to the CD24 expressing cancer cells. To the best of our knowledge, this study is the first report to identify an scFv antibody against CD24, using the phage display method. [ABSTRACT FROM AUTHOR]

Highlights: [•] The highest antileishmanial activity was seen for Ag NPs. [•] UV and IR light increased antileishmanial properties in all nanoparticles. [•] Nanoparticles had same cytotoxicity for both macrophages and promastigotes. [ABSTRACT FROM AUTHOR]

Highlights: [•] TG coating led to decreased nanoparticle cytotoxicity by decrease in ROS generation in Balb/c mice skin cells. [•] UV or IR light led to increased cytotoxicity of coated nanoparticles by increase in ROS generation. [•] Metal nanoparticles were more toxic than metal oxide nanoparticles. [ABSTRACT FROM AUTHOR]

Abstract: GIRK2 is the primary post-synaptic effector of opioids in the CNS that may contribute to the sex differences or age dependency in opioid analgesia. In the present study, we investigated the differential developmental and hormonal regulation of GIRK2 gene transcription in rats. Male and female Wistar rats were divided into infant or adult intact, gonadectomized, sham and gonadectomy+testosterone groups. Semi-quantitative RT-PCR was used to determine the levels of GIRK2 gene expression in spinal cord and brain. Our results showed that in male spinal cord/brain, the gene transcription of the infant group did not differ from expression levels of this gene in the gonadectomized or adult intact groups. Castration of males significantly decreased the expression levels of the GIRK2 gene. Testosterone replacement in castrated rats did not raise GIRK2 gene expression completely to the level of shams. In females however, a greater expression of GIRK2 gene expression was found in adult intact or gonadectomized rats than in the infant groups. Ovariectomy failed to alter GIRK2 mRNA levels significantly. No significant sex differences were observed in GIRK2 gene transcription between intact, sham and infant groups, but gonadectomy produced sex differences in GIRK2 gene transcription. The results strongly demonstrate the differential developmental and hormonal regulation of GIRK2 in the rat CNS. [Copyright &y& Elsevier]

Abstract: Locus coeruleus (LC) plays a key role in opioid dependence and withdrawal. Chronic morphine administration induces neurochemical adaptations in the noradrenergic system. The nature of signal responsible for opiate-induced adaptations of noradrenergic neurons in LC is not well defined. Neurotrophins-signaling pathways such as brain derived neurotrophic factor (BDNF) and Neurotrophin-3 (NT-3) play a key role for regulating the noradrenergic response of LC neurons to opiates. The nucleus paragigantocellularis (PGi) is one of the two major afferents to LC. The present study was designed to evaluate the expression of BDNF and NT-3 in the context of opiate dependence and withdrawal in PGi. Such data are important because they could reveal the role of PGi as an additional source of BDNF and NT-3 in the neurochemical plasticity of LC neurons. Opiate dependence was induced by a progressive intraperitoneal treatment of morphine. In morphine dependent group PGi nucleus was extracted for gene expression assay 6h after the last injection of morphine. In spontaneous withdrawal, rats received the same chronic treatment as morphine group. PGi was extracted for gene expression assay 24, 48 and 72h after the last injection of morphine. PGi nucleus was assayed for the expression of BDNF and NT-3 using semi-quantitative RT-PCR normalized to β-actin gene expression. Results showed that chronic administration of morphine significantly increased BDNF and NT-3 gene expression in PGi. In spontaneous withdrawal, BDNF/NT-3 genes expression were high in comparison to control group. It seems that BDNF/NT-3 -signaling pathway originating from PGi is essential for opiate-induced adaptations of the LC neurons. [Copyright &y& Elsevier]

Abstract: It is previously reported that the HPA axis plays role in the inhibitory effect of pain on tolerance development to analgesic effect of opioids. The present study was designed to investigate whether the chronic co-administration of dexamethasone as a glucocorticoid is also able to prevent or reverse analgesic tolerance to morphine and to compare the expression of Gαi/o and Gβ subunits of G proteins in the context of chronic dexamethasone, development of morphine tolerance and their combination. Analgesic tolerance to morphine was induced by chronic intraperitoneally (i.p.) administration of morphine 20 mg/kg to male Wistar rats weighing 200–240 g within 4 consecutive days and analgesia was assessed using tail-flick test. Chronic dexamethasone was applied using 4 daily i.p. injections. Lumbar spinal tissues were assayed for the expression of Gαi/o and Gβ proteins using “semiquantitative PCR” normalized to beta-actin gene expression. Results showed that chronic administration of dexamethasone could reduce and reverse the development of tolerance in rats that received chronic i.p. injections of morphine. Chronic administration of dexamethasone significantly increased the expression of Gαi/o, while chronic administration of morphine did not change its expression. The expression of Gβ, however, was increased after the chronic administration of morphine, but did not change after the administration of chronic dexamethasone. None of these increases were observed when morphine and dexamethasone were co-administered. We conclude that the development of tolerance to analgesic effect of morphine could be prevented and reversed by dexamethasone co-administration. The increase in Gαi/o genes expression produced by chronic dexamethasone may facilitate the opioid signaling pathway and compensate for morphine-induced tolerance. [Copyright &y& Elsevier]

Ureteral obstruction with subsequent hydronephrosis is clinically common. Most cases are identified and diagnosed in the perinatal period. The diagnosis of ureteropelvic junction obstruction (UPJO) implies a functionally significant impairment of the urinary transportation from the renal pelvis to the ureter. Although most cases are probably congenital in origin, they can clinically remain hidden until much later in life. UPJO is usually considered an isolated event. Recently, we have evaluated a father and his 3 sons, all of whom had UPJO. This study reports a missense mutation of threonine 386, which was replaced with alanine in Wilms'' tumor genes. We suggest that UPJO might not necessarily be sporadic and other family members might have a similar problem. [Copyright &y& Elsevier]

The healing process of non-healing and full-thickness wounds is currently facing some serious challenges. In such ulcers, losing a large part of skin causes a chronic infection due to the entrance of various pathogens in the wound bed. Moreover, poor vascularization, uncontrolled inflammation, and delayed re-epithelialization increase the healing time in patients suffering from such wounds. In this light, tissue engineering provides a wide range of strategies using a variety of biomaterials, biofactors and stem cells to decrease the healing time and restore the function of the damaged site. A suitable wound healing agent should possess some critical parameters such as inducing re-epithelialization, anti-inflammatory and anti-bacterial properties, and angiogenic capability. The Lacto-n-Neotetraose (LNnT) with chemical formula C26H45NO21 is an oligosaccharide present in human milk and soluble antigens extracted from Schistosoma mansoni eggs. It is reported that LNnT induces type 2 immune response (Th2 immunity). Th2 immunity promotes re-epithelialization, angiogenesis and wound contraction by recruiting the cells which produce Th2-related cytokines. Moreover, LNnT shows some special characteristics such as angiogenic capability, anti-inflammatory, and anti-bacterial effects which can address the mentioned challenges in the treatment of non-healing and full-thickness wounds. Here, we hypothesize that utilizing LNnT is an appropriate biofactor which would improve the healing process in full-thickness and non-healing wounds. [ABSTRACT FROM AUTHOR]

• The immune-dominant epitopes could act as efficient alternatives for whole immunogenic antigens. • The designed HHP construct contains immune-dominant epitopes of HBsAg , HCVcp , and VPs. • The HHP antigen has enhanced DC maturation capacity compared to single subunit counterparts. • The DCs induced by HHP antigen are able to trigger T-cell activation and proliferation. • The designed HHP antigen may be utilized as a vaccine candidate against hepatitis and polio viruses. Design and application of epitope-based polyvalent vaccines have recently garnered attention as an efficient alternative for conventional vaccines. We previously have reported the in silico design of HHP antigen which encompasses the immune-dominant epitopes of Hepatitis B surface antigen (HBsAg), Hepatitis C core protein (HCVcp) and Poliovirus viral proteins (VP s). It has been shown that the HHP has desirable conformation to expose the epitopes, high antigenicity and other desired physicochemical and immunological properties. To confirm the accuracy of these predictions, the ex-vivo immunogenicity of the HHP was assessed. The HHP gene was chemically synthesized in pET28a and expressed in E. coli (BL21). The expressed protein was purified and its immunological potency was evaluated on dendritic cells (DCs) as antigen presenting cells (APCs). Functional analysis was assessed in co-cultivation of autologous T-cells with matured DCs (mDCs). T-cell activation, proliferation and cytokines secretion were evaluated using flowcytometry and ELISA methods. Our results indicated that the HHP could induce the DC maturation. The mDCs were able to trigger T-cell activation and proliferation. In silico design and ex-vivo confirmation of immunological potential could pave the way to introduce efficient immunogens for further analysis. The ability of HHP in DC maturation and T-cell activation makes it an amenable vaccine candidate for further in-vivo studies. [ABSTRACT FROM AUTHOR]

• Expression of DHFR and PTR1 genes. • Real-time PCR method for gene expression. • The effect of OLE on the expression of genes in leishmania major. Among neglected tropical diseases (NTD) following malaria, leishmaniosis has claimed the largest mortality rate (Georgiadou, Makaritsis, and Dalekos 2015). None of the current anti-leishmanial drugs can be considered ideal (Minodier and Parola 2007; Mohapatra 2014). The dihydrofolate reductase- thymidylate synthase (DHFR-TS) leads to DNA synthesis in the parasite and its inhibition causes the death of the parasite. Meanwhile, pteridine reductase1 (PTR1) enzyme, because of having a similar role to DHFR in the parasite, causes by-pass of DHFR enzyme inhibition thereby resulting an incidence of drug resistance in the parasite (Nare et al. 1997b). Considering the current problems regarding leishmaniosis treatment, developing new drugs with less toxicity and greater efficacy plus lower costs has changed into a necessity. The present study examined the effect of olive leaf extract (OLE) at concentrations 6.25, 12.5, 25, 50, and 100 µg/ml on the expression of DHFR and PTR1 genes in Leishmania major parasite. The parasite samples were incubated in RPMI 1640 culture medium in the presence of five mentioned concentrations of OLE. A quantitative real-time PCR technique was used to analyze the DHFR and PTR1 gene expressions. The results revealed that the expression of DHFR gene at 6.25 µg/ml extract increased significantly (p <0.05), while it diminished significantly at 25 µg/ml. At other studied concentrations, DHFR gene showed underexpression compared to the control group, which was not statistically significant (p >0.05). The expression of PTR1 gene at concentrations 6.25, 50, and 100 µg/ml of the olive leaf extract increased significantly (p <0.05). The expression of PTR1 gene at 12.5 µg/ml concentration decreased compared to the control group, while it increased at 25 µg/ml, though the changes in the gene expression at these two concentrations were not statistically significant (p >0.05). The findings of the present study suggest that OLE can reduce DHFR expression, but its effect of reducing PTR1 expression was not very notable, with its influence being greater on DHFR underexpression rather than PTR1. [ABSTRACT FROM AUTHOR]

At present, there are no efficacious vaccines or effective drugs against leishmaniasis; therefore new and innovative control methods are urgently required. One way to achieve this important goal is through using reverse genetic engineering to evaluate important enzymes, proteins and macromolecules. One of the most important enzymes for Glycosylphosphatidylinositol (GPI) biosynthetic pathways is GlcNAc-PI-de-N-acetylase ( GPI12 ). The molecular constructs were cloned in Escherichia coli strain Top 10 and confirmed by molecular methods and were transfected by electroporation into L eishmania major . We demonstrated that two alleles of the GPI12 gene in L. major were successfully removed and enabling the generation of a null mutant, which supports the idea that GPI12 is not an essential gene for the growth and survival of Leishmania and the homozygous knockouts of Leishmania are able to survive. We were able to produce a mutant parasite that caused no damaged to the host. Further investigations are essential to check the safety profile in laboratory animals. [ABSTRACT FROM AUTHOR]

Insulin production under the stringent control is the main issue in gene-based therapeutic strategies directed to type 1 diabetes. As a novel approach, inducible promoters may provide a promising tool for this purpose. In this study, we hypothesize that this control may be achieved via a promoter derived from the heat shock multigene family, Hsp70 A1A, which is inducible at 42°C. To yield mature insulin in transfected fibroblasts (3T3/NIH), a recombinant human insulin gene consisting of sequences corresponding to furin cleavable sites was fused to the promoter. Heat-stimulated cells initiated to release biologically active insulin within 30 min with a ten-fold increase after 24 h. The role of upstream regulatory elements of the promoter on its activity in heat stress conditions was examined. No significant difference between the activity of the minimal and full-length promoters was observed. This promoter exhibited low basal expression in non-inducing conditions. Results indicate that this promoter is responsive to a heat induction after approximately 30 min which causes an efficient insulin production over a relatively short period of time. These potential features of this promoter may provide an insight to control the insulin production in vivo upon an external and physical stimulation. [ABSTRACT FROM AUTHOR]

Abstract: This study has identified and characterized the structure of locus D-A, a noncoding short tandem repeat (STR) region, also known as locus 1–2, in Iranian Entamoeba dispar isolates. This polymorphic locus has been shown to be potentially useful in investigating the molecular epidemiology of Entamoeba histolytica and E. dispar. The genetic polymorphisms in locus D-A in 28 isolates of E. dispar from three different geographic regions of Iran were distinguished using PCR and sequencing, and the results were compared with the E. dispar gene sequences available in GenBank. In all microscopy-positive E. histolytica/E. dispar samples, PCR with species-specific primers was used to amplify a 477–531bp product, identifying the samples that had E. dispar. Analysis of the sequences revealed a remarkable degree of genetic diversity with regard to size, number and organization of the repeat units among the E. dispar isolates. The sequenced products showed 12 novel E. dispar genotypes, which have been submitted to the GenBank/EMBL/DDBJ database under accession numbers AB354125–AB354136 AB354125 AB354126 AB354127 AB354128 AB354129 AB354130 AB354131 AB354132 AB354133 AB354134 AB354135 AB354136 . [Copyright &y& Elsevier]

Safe and efficient delivery of microRNA (miRNA) molecules is essential for their successful transition from research to the clinic setting. In the present study, we have used a bile acid, deoxycholic acid (DA), to modify 1.8 kDa branched polyethylenimine (bPEI 1.8) and subsequently investigated gene delivery features of the resultant conjugates (PEI-DA n). We found significant differences between the PEI-DA n conjugates and conventional bPEIs with respect to miRNA condensation ability, buffering capacity, cellular uptake, and intracellular gene release behavior in endothelial cells (ECs) isolated from human umbilical vein (HUVECs). Changes in the conjugation degree greatly influenced the transfection performance of the PEI-DA n conjugates with respect to miRNA condensation and decondensation properties as well as cellular uptake behavior. The PEI-DA 3 conjugates could significantly enhance the expression level of miRNA-210 in HUVECs. The overexpressed miRNA-210, in turn, markedly downregulated the expression levels of Efna3 and Ptp1b as well as led to a substantial rise in HUVECs' migration rate in a wound healing assay. Collectively, our results have demonstrated that PEI-DA 3 conjugates facilitate the formation of stable nanocomplexes that are loose enough to release miRNAs into the cytosol. The free bioavailable miRNAs, in turn, result in efficient gene silencing comparable to bPEI 25 as well as Lipofectamine RNAiMAX. [ABSTRACT FROM AUTHOR]

Highlights • Urinary stones are the third most commonly reported urinary tract disease. • Oxalic acid is highly oxidized organic compounds, which found in dietary and produced the intestinal microflora. • In this study, the Oxalyl CoA decarboxylase gene contains an-histidin tag was cloned in pET28a (+) and expressed in Escherichia coli BL21 (DE3). • The antibody against Oxalyl CoA decarboxylase protein was used in ELISA assay. • This study suggests that antibody against Oxalyl-CoA decarboxylase protein is useful to measure oxalyl CoA decarboxylase enzymes in patients. Abstract Urinary stones are the third most commonly reported urinary tract disease. Kidney stones are one of the most common types of urinary stones that most of them (70–80%) are calcium oxalate. Oxalic acid is highly oxidized organic compounds, which found in dietary and produced by the intestinal microflora. Oxalyl CoA decarboxylase is a key enzyme and plays an important role in oxalate degradation. In this study, the Oxalyl CoA decarboxylase gene which contains an –histidine tag was cloned in pET 28a (+) and expressed in Escherichia coli BL21 (DE3). The purified Oxalyl CoA decarboxylase protein was injected into rabbit for immunization. The antibody against Oxalyl CoA decarboxylase protein was used in ELISA assy. eventually, this ELISA system was used for patients with calcium oxalate kidney stones. ELISA analysis of serum samples of patient white calcium oxalate kidney stones showed that 88.8% of patient was lacking in antibody against Oxalyl CoA decarboxylase. This study suggests that antibodies against oxalyl-co-decoxylase proteins are useful in the detection of urinary tract stones and It can be used to measure oxalyl CoAdecoxylase enzymes in a simple method. [ABSTRACT FROM AUTHOR]

After half a century, Trichinella infection has been reported in humans following ingestion of wild boar meat hunted in Northern Iran. We have performed a cross-sectional study to evaluate the prevalence of Trichinella spp. infections in wild boar and prevalence of anti- Trichinella antibodies among at-risk individuals for the first time in Iran. Muscle and sera samples were collected from 79 wild boars and 364 at-risk individuals. Trichinella infection has been investigated by artificial digestion and molecular identification (in wild boar muscles) and by serology (in humans). Trichinella larvae were isolated from three wild boars (3.7%; 95% CI, 2.9–4.3). The isolated larvae were identified as T. britovi using CO1 and 5S rRNA gene primers amplification. The percent identity regarding to 5S rRNA gene (98.7–100%) and divergence (0–1.9%) further confirmed the isolates as T. britovi . Of the 364 participants, anti- Trichinella IgG were detected in 8 (2.2%; CI 95%, 1.9–2.4). Risk factors associated with Trichinella infection seropositivity in humans, were hunter being (OR, 13.5; 95% CI, 3.1–59.4; P = 0.003) and high consumption (more than 7 time in a year) of wild boar meat (OR, 17.5; 95% CI, 3.2–93.6; P < 0.001). In conclusion, results of this study emphasized that consumption of wild boar meat could be important source of human trichinellosis as a completely neglected infection disease in Iran. We suggest that the additional studies should be performed in different parts of Iran to further clarify the prevalence of trichinellosis in wild animals to guide the development of appropriate public health interventions. [ABSTRACT FROM AUTHOR]

Aim Human papillomaviruses (HPV) have frequently been detected in colorectal cancer tumor samples, and may play a role in the pathogenesis of colorectal cancer. This study was designed to investigate the presence of DNA and RNA for the high-risk HPV genotypes 16 and 18 in samples of colorectal cancer tumors and adjacent normal tissues. We also investigated the expression of proapoptotic genes in HPV-positive colorectal tumors compared to normal tissue samples. Methods Samples of tumoral and adjacent normal tissues were fresh-frozen, and HPV DNA was identified by nested and semiquantitative PCR. Real time PCR was used to quantitatively compare the expression of HPV-18 E6 and nine proapoptotic genes in HPV-positive tumors and samples of adjacent normal tissue. Results HPV-16 DNA was found in 10.5% of the tumor samples, and HPV-18 DNA was found in 23.6% of the samples. Real time PCR results showed lower expression of the E6 gene in HPV-positive tumors than in adjacent normal tissue. The expression of two proapoptotic genes, FAS and DR5 , was significantly lower in tumor samples than in adjacent normal tissues. Conclusions HPV infection, especially HPV-18, may play a role in colorectal cancer tumorigenesis by downregulating death receptor genes and interfering with the extrinsic pathway of apoptosis. [ABSTRACT FROM AUTHOR]

The aim of this study was to find the peptide ligands to inhibit Candida albicans secreted aspartyl proteinase 2 (Sap2). First, a ligand library, containing 300 different peptides, was constructed, and their interaction with Sap2 was separately calculated by molecular dynamic software. Second, 10 peptide ligands with the lowest intermolecular energy were selected. Then, triangular gold nanoparticles were synthesized, and separately conjugated with the peptide ligands. After synthesis, antifungal property and Sap inactivation of conjugated triangular gold nanoparticles, peptide ligands, and naked triangular gold nanoparticle were separately assessed, against thirty clinical isolates of C. albicans. In this study, we measured the uptake of conjugated and naked nanoparticles by atomic adsorption spectroscopy. This study showed that naked triangular gold nanoparticle and all conjugated triangular gold nanoparticles had high antifungal activity, but no peptide ligands had such activity. Of 300 peptide ligands, the peptide containing N-Cys-Lys-Lys-Arg-Met-Met-Lys-Ser-Met-Cys-C and its conjugate had the highest capability to inhibit Sap. Moreover, the uptake assay demonstrated that triangular gold nanoparticles conjugated with the peptide ligand had the highest uptake. [ABSTRACT FROM AUTHOR]

Abstract: Toxoplasmosis diagnosis constitutes an important measure for disease prevention and control. In this paper, a newly described DNA amplification technique, loop-mediated isothermal amplification (LAMP), and nested-PCR targeting the repeated element (RE) and B1 gene, were compared to each other for the detection of Toxoplasma gondii DNA in blood samples of children with leukaemia. One hundred ten blood samples from these patients were analyzed by LAMP and nested-PCR. Out of 50 seropositive samples (IgM+, IgG+), positive results were obtained with 92% and 86% on RE, B1-LAMP and 82% and 68% on RE, B1-nested PCR analyses, respectively. Of the 50 seronegative samples, three, two and one samples were detected positive by RE-LAMP, B1-LAMP and RE-nested PCR assays, respectively, while none were detected positive by B1-nested PCR. None of the 10 IgM−, IgG+ samples was detected positive after testing LAMP and nested-PCR assays in duplicate. This is the first report of a study in which the LAMP method was applied with high sensitivity and efficacy for the diagnosis of T. gonii in blood samples of children with leukaemia. [Copyright &y& Elsevier]

Abstract: Stem cell therapy is a promising approach for treatment of degenerative retinal disorders such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD). In this study, human mesenchymal stem cells (MSCs) were isolated from the trabecular meshwork (TM), the major functional tissue of the anterior chamber angle in the eye, were characterized and differentiated into photoreceptor cells on amniotic membrane (AM). After isolation of trabecular meshwork and culture of the stromal segment of this tissue, fibroblast-like cells (CD105+, CD90+, CD44+, CD166+ cells) capable of differentiation toward mesenchymal and photoreceptor lineages were obtained. The isolated cells were seeded on amniotic membrane and were treated with induction medium. Immunocytochemistry and quantitative real time RT-PCR (qPCR) were used to detect expression of photoreceptor genes such as rhodopsin, recoverin, CRX, and peripherin; and the bipolar cell marker protein kinase C alpha (PKC-alpha). As a result, immunocytochemistry revealed that the differentiated TMMSCs expressed rhodopsin, CRX and PKC proteins. qPCR showed the expression of rhodopsin (rod like photoreceptor-specific marker), and CRX genes were significantly higher in TMMSCs differentiated on AM than those differentiated on tissue culture polystyrene (TCPS). In conclusion, our findings suggested that a combination of TMMSCs (as a new source) and basement membrane support from AM might be a suitable source of cells for subretinal transplantation in regenerative therapy for retinal disorders such as AMD and RP. [Copyright &y& Elsevier]

Abstract: Clinically infected dogs have been identified as the main reservoir hosts of visceral leishmaniasis (VL) caused by Leishmania infantum in the Mediterranean region. The objective of this study was to determine the potential of asymptomatic infected dogs compared with symptomatic ones as a source of L. infantum infection to golden hamster. For this purpose, anti-Leishmania antibodies were detected with direct agglutination test (DAT) in 13 symptomatic (7 seropositive=≥1:320) and 53 asymptomatic (9 seropositive=≥1:320 and 44 seronegative= <1:320) ownership dogs. DNA of Leishmania sp. was extracted from skin and peripheral blood tissues of each dog and tested by PCR. Sixty-six Syrian golden hamsters (Mesocricetus auratus) were used for the determination of infectivity and pathogenicity of L. infantum, isolated from the dogs. We used the internal transcribed spacer 2 (ITS 2) rDNA sequence analysis. The results showed that 22 and 11 out of 66 inoculated golden hamsters were positive by PCR and parasitological examinations, respectively. From 22 PCR positive hamsters, 17 were related to asymptomatic dogs and 5 were from symptomatic ones. There was no significant difference between symptomatic and asymptomatic dogs in producing Leishmania infection in the susceptible animal model (P =0.66). Smears and cultures of 5 dogs from 13 symptomatic dogs (38.5%) and 6 dogs from 53 asymptomatic ones (11.3%) were found to be positive at parasitological examination. All the L. infantum isolates from symptomatic and asymptomatic dogs were similar in sequencing. In conclusion, asymptomatic infected dogs as well as symptomatic ones can harbor L. infantum in their blood and skins which are virulent and infectious for inoculated golden hamster. [Copyright &y& Elsevier]

Abstract: There are some reports regarding the inhibitory effect of pain on tolerance development to analgesic effect of opioids. The present study was designed to investigate whether the chronic formalin induced pain is able to reverse analgesic tolerance to morphine and to evaluate the expression of Gαi/o and Gβ subunits of G proteins in the context of chronic pain, development of morphine tolerance and their combination. Morphine tolerance was induced by chronic systemic (intraperitoneally, i.p.) or spinal (intrathecally, i.t.) administration of morphine to male Wistar rats weighing 200–240g and analgesia was assessed using tail flick test. Chronic pain was induced by 4 daily intraplantar injections of 50μl of 5% formalin. Lumbar spinal tissues were assayed for the expression of Gαi/o and Gβ proteins using “semiquantitative PCR” normalized to beta-actin gene expression. Results showed that chronic formalin induced pain could reduce and reverse the development of tolerance in rats that had received chronic (i.p. or i.t.) administration of morphine. Chronic administration of morphine did not change Gαi/o gene expression, while chronic pain significantly increased its expression. The expression of Gβ, however, was increased after the chronic administration of morphine, but did not change after the induction of chronic pain. None of these increases were observed when morphine and formalin were administered at the same time. Due to synchronous development of morphine tolerance and changes in expression of Gβ, it may be concluded that the development of tolerance to analgesic effect of morphine is partially mediated by increase in Gβ gene expression. The increase in Gαi/o genes expression produced by chronic pain may facilitate the opioid signaling pathway and compensate for morphine-induced tolerance. [Copyright &y& Elsevier]

Colorectal cancer (CRC) is the fourth most common cause of cancer and mortality worldwide and is the third most common cancer in men and women. Surgery, radiotherapy, and chemotherapy are conventionally used for the treatment of colorectal cancer. However, these methods are associated with various side effects on normal cells. Thus, new studies are moving towards more effective and non-invasive methods for treatment of colorectal cancer. Targeted therapy of CRC is a promising new approach to enhance the efficiency and decrease the toxicity of the treatment. In targeted therapy of CRC, antibody fragments can directly inhibit tumor cell growth and proliferation. They also can act as an ideal carrier for targeted delivery of anticancer drugs. In the present study, the structure and function of different formats of antibody fragments, immune-targeted therapy of CRC using antibody fragments will be discussed. Targeted therapy for CRC treatment. Cancer cells express different receptors and antigens that can be targeted by antibody fragments. Many of these receptors promote the invasion, survival, angiogenesis, proliferation, metastasis and development and inhibit apoptosis pathways that with antibody fragments can inhibit the effects of these antigens and CRC. Unlabelled Image [ABSTRACT FROM AUTHOR]

Abstract: The aim of this study was to determine the prevalence, variability with host age, and the genotypes of species of Cryptosporidium in cattle from 15 dairy farms in Qazvin province, Iran. Fecal samples, collected from 272 cattle during May 2006 to December 2007, were characterized microscopically. Oocysts from 51 positive samples were analyzed using PCR assay of 18S SSU rRNA, restriction fragment length polymorphism (RFLP) and sequencing. We identified 72.6% of the positive samples as Cryptosporidium parvum, 17.7% as Cryptosporidium andersoni, 7.8% as Cryptosporidium bovis and 1.9% as a novel genotype of C. parvum possessing a single mutation on MboII restriction. An infection rate of 19.5% of C. parvum among 174 pre-weaned calves was significantly higher than the 3.1% among 98 post-weaned calves (P <0.0006). This is the first report of C. bovis and the new subgenotype of C. parvum in Iranian cattle. [Copyright &y& Elsevier]