fattah sotoodehnejadnematalahi

Glioblastoma (GBM) is an aggressive and malignant brain cancer with the highest mortality and low survival rates. To discover a more specific and efficient treatment for GBM, we synthesized and examined the cytotoxic effect of arazyme-interleukin-13 (Ara-IL13) fusion protein on GBM cells.

Experimental approach: 

At first, the araA-IL13 chimeric gene in the pET28a (+) vector was designed and synthesized. After transformation into Escherichia coli BL21 (DE3), the chimeric gene was verified by colony polymerase chain reaction. Expression optimization and purification of the AraA-IL13 fusion protein was performed and subsequently evaluated by 10% SDS-PAGE. The protein was purified and concentrated using the Amicon® Ultra- 15 centrifugal filter unit. The presence of AraA-IL13 was investigated by the western blotting technique. The enzyme was evaluated for proteolytic activity after purification on skim milk agar. The cytotoxic effect of the AraA-IL13 fusion protein was evaluated by MTT assay on U251 and T98G cell lines in vitro.

The chimeric protein had no proteolytic activity on skim milk agar despite high expression. Furthermore, no cytotoxic effect of this fusion protein (up to 400 μg/mL) was observed on the U251 and T98G cell lines.

Conclusion and implications: 

The lack of proteolytic activity and cytotoxic effect of AraA-IL13 may be due to the disruption of the three-dimensional structure of the protein or the large structure of the arazyme coupled with the ligand and the lack of proper folding of the arazyme to make the active site of the enzyme inaccessible.

Extracellular vesicles (EVs) are nano- to micron-sized vesicles with the ability to transport bioactive cargos. All types of cells have the ability to release EVs, which have been demonstrated to be involved in a number of essential cell functions. There are various methods for isolating EVs, each with advantages and disadvantages. The purpose of this study was to invistigate at the non-ultra centrifugation method and ultra-centrifugation method for isolating EVs.

Clostridium perfringens ATCC13124 was used in this experimental study. Following culture, EVs were extracted using two methods Non-Ultra method and Ultra-method. To examine chemical characteristics, the EV protein concentration was measured using a NanoDrop device, and the EV protein pattern was identified using SDS-PAGE. Transmission electron microscopy (TEM) was utilized to examine the EVs' physical properties.

Our results showed that the EVs isolated by the Ultra-method had a higher protein content compared to the Non-Ultra method (3.17 and 1.46 mg/ml, respectively).The Ultra-method isolated more and larger EVs compared to the Non-Ultra method. Also, protein patterns of the EVs by SDS-PAGE method were similar in both methods.

The present study showed that the Ultra-method can be a more efficient and cost-effective way for isolation EVs from Clostridium perfringens than the Non-Ultra method.

COVID-19, an infectious viral disease caused by severe acute respiratory syndrome (SARS-CoV-2) with more than 260 million infections as of December 2019, is a serious threat to the health and economy of human societies. Designing and producing a suitable vaccine can Reduce the incidence of this disease. Therefore, it is very important to find effective and safe neutralizing antibodies and vaccines for COVID-19. The RDB section of the spike protein is a suitable option for the production of subunit vaccines and neutralizing antibodies. This research aims to introduce the RBD section as a vaccine candidate.

The RBD was recombinantly expressed in two hosts, Escherichia coli (E.coli) and insect cells, and its production rate was compared. Using the western blotting method, protein production was confirmed. To evaluate the function of the protein expressed in two prokaryotic and eukaryotic hosts, using the serum of patients recovered from COVID-19 (wild type and delta), the ELISA method was used.

Despite the higher production of recombinant protein by E.coli, the affinity of the antibodies in the serum of the patients to the protein expressed in the insect cell was higher.

In this study, the RBD was expressed in two different hosts. The results show that RBD is mentioned as a vaccine candidate. Homozygous in the insect cell preserves the biological activity of the protein by carrying out the process of post-translational changes such as glycosylation.

Given the global epidemic of COVID-19, it is important to design a vaccine for prevention. The virus belongs to the beta-coronavirus family and forms appendages on the surface of the glycoprotein spike virus membrane. Studies on SARS-CoV-1 and related MERS-CoV vaccines have shown that the spike protein on the virus surface is a suitable target for the vaccine. In this experimental study, we compare the recombinant fragment of spike protein (rfsp) expressed in the eukaryotic host CHO-K1 cell and prokaryotic E. coli in terms of immunogenicity, neutralizing activity, and epitopes recognition Similar to the virus strain and the ability to bind to the serum of improved patients, in two types of alpha and delta variants. The results showed that both rfSP proteins are a potential new antigen candidate for the development of the Covid 19 vaccine, but the CHO cell maintains the biological activity of the protein by performing post-translational modification processes such as glycolysis. This increases the present likelihood of developing virus-like epitopes and increases the titer of rfsp-specific antibodies in the serum of immunized mice. Therefore, priority is given to rfsp expressed in CHO cells to evaluate vaccine efficacy.

Viruses are obligate parasites and are completely dependent on host cells for survival and replication. RBK cell line developed and introduced by Razi Vaccine and Serum Research Institute, has been successfully established as a continuous cell line over successive passages.Viruses are obligate parasites that completely rely on host cells for survival and replication. Razi Bovine Kidney (RBK) cell line was introduced and developed by the Razi Vaccine and Serum Research Institute. It has been successfully established as a continuous cell line over successive passages. As demonstrated in this experimental study, the RBK cells have shown suitable sensitivity to certain viruses, including Bovine Herpesvirus-1 (BoHV-1) virus. In the present research, the RBK cell line characteristics were analyzed using molecular and karyotype methods and growth specifications. Cloning of the RBK cell line was performed using the limited dilution method, and each cell clone was quantitatively and qualitatively characterized. Four cell clones were compared based on their sensitivity to the BoHV-1 virus. Then, the RBK-D5 clone was selected as the most appropriate cell line for further studies. The RBK-D5 was subjected to tests for identity, chromosomal analysis, and doubling time. In the end, the origin of this cell line was confirmed by the PCR method. It was observed that the cell line exhibited karyotype diversity due to aneuploidy, which can be responsible for the procreation of chromosomal instability. This diversity represents chromosomal changes in the continuous cell line that carries the characteristic of an immortalized cell line. The RBK-D5 was found to be more sensitive to the BOHV-1 virus. Surprisingly, its titer was evaluated at 108.5 CCID50/ml. The obtained results suggested that the RBK cell line is suitable for the BoHV-1 virus and can be useful for virus detection, propagation, and quality control or viral titration.

Kidney diseases are an important medical problem worldwide. Since there are limited treatment options for damaged kidneys, stem cell therapy has become an alternative treatment. The aim of present study is to investigate effect of culture medium obtained from mesenchymal stem cells (MSCs-CM) of newborn mice kidneys in percentages of 10, 30 and 50 on differentiation of embryonic stem cells towards kidney epithelial cells. Mesenchymal stem cells were isolated from kidneys of newborn mice, passaged and propagated. Determination of the identity of cells was done using flow cytometry and checking the expression of surface markers CD105, CD29, CD90. In third passage of extracted cells, supernatant culture medium was collected, hESC were cultured and multiplied in the complete culture medium of cells and the differentiation of hES cells into progenitor cells was investigated. The expression of PAX2, ZO1 and CK18 genes was investigated using RT-PCR, expression of CD133, CD24 and CD44 surface markers was investigated using flow cytometry. Flow cytometry results confirmed the mesenchymal nature of the cells. The results of differentiation of hESCs showed that expression of PAX2, ZO1 and CK18 genes increased significantly (p<0.05) in the groups containing supernatant. The results of flow cytometry show an increase in expression of CD133 and CD24 markers in groups containing CM and the expression of CD44 marker in the group containing 50% CM, compared to control group. In general, results showed supernatant culture process of cells has a positive effect on inducing differentiation of human embryonic stem cells into kidney progenitor cells.

Since December 2019, the world has been grappling with an ongoing global COVID-19 pandemic. Various virus variants have emerged over the past two years, each posing a greater threat than its predecessors. The recent appearance of the omicron variant (B.1.1.529) has raised significant alarm within the field of epidemiology due to its highly contagious nature and rapid transmission rate. The omicron variant possessed mutations in the key receptor-binding domain (RBD) region, the S region, and these modifications have shown a notable impact on the strain's susceptibility to neutralizing antibodies. Developing safe and efficient vaccines to prevent a future severe acute respiratory outbreak of coronavirus syndrome 2 (SARS-CoV-2) is significant. Viral surface spike proteins are ideal targets for vaccines. This study aimed to find a multi-subunit chimeric vaccine. After conducting bioinformatics analysis, the recombinant spike (RS) protein of SARS-CoV-2 was deliberately designed and subsequently produced using E. coli expression systems. The immunogenicity of RS and neutralizing antibody responses were evaluated on immunized BALB/c mice. There was a significant difference in antibody titers between RS-immunized mice and control groups. The endpoint of the serum antibody titer of mice immunized with our chimeric protein was 2.5 times higher than that of the negative control. The chimeric construct could present multiple antigens simultaneously, influentially affecting immunization. Sera from mice vaccinated by RS could recognize the SARS-CoV-2 virus and neutralize antibodies. Our chimeric peptide could bind to antibodies in the serum of patients infected with different serotypes of the SARS-CoV-2 virus, such as alpha, delta, and omicron variants. The results indicated that the RS protein would be a potential novel antigenic candidate for subunit vaccine development and could be used as a useful alternative to generate diagnostic serological tests for SARS-CoV-2 infection.

 Defensive barrier is important in preventing infection and disease progression caused by E.coli O157:H7. Among beneficial effects of probiotics, immune system modulation by producing cytokines is one of the most common reported benefits. The aim of this study was to investigate the Lactobacillus acidophilus probiotic effect on il10 cytokine gene expression in Zebrafish infected with E. coli O157:H7 as a laboratory model.

 The 300 pieces of fish were grouped equally; control group (C1A1) received basic diet, group (B1A1) received basic diet and faced E.coli O157:H7, treatment 1 group (T1A1) received basic diet containing L. acidophilus and no exposure to E. coli O157:H7, treatment 2 group (T2A2) received basic diet containing L. acidophilus and exposed to E. coli O157:H7. During the 30-day period of testing, intestinal tissue samples were taken on days 15, 27 and 30 for il10 gene expression and bacterial colonization studies.

 The findings indicated bacterial colonization in fish intestines. Also, the expression of il10 gene in the experimental groups decreased after exposure to E. coli O157:H7 (P<0.01). This decrement was less with the regulation of il10 in the groups fed with L. acidophilus compared to the control group. In Zebrafish fed with L. acidophilus, the expression of the il10 gene increased gradually (day15, day27 and 30; P<0. 01).

 Overall, the results showed that the L. acidophilus was able to increase il10 gene expression, boost the immune system in Zebrafish as a laboratory model. Therefore, L. acidophilus La5 probiotic can be used as a possible useful candidate against E. coli O157:H7.

Background &

In today's societies, breast cancer has significantly threatened women's health (1). Successful cancer treatment can also destroy tumor germ cells (2, 3). Treatment methods based on immune checkpoint molecules are considered (4). PD-L1 overexpression suppresses the immune response to cancer cells and causes epithelial-to-mesenchymal transition (EMT) and cancer stem cell (CSC) phenotypes, metastasis and chemical resistance (5). Overexpression of PD-L1 and MUC1 is observed in approximately 90 TNBCs. In this present study, we investigated the inhibitory effect of miR-143-3p on MUC1 expression as an oncogene that has the most impact on PDL-1 overexpression in BC cell lines MCF-7 (less aggressive BC), MDA‐MB‐231, and BT-549 (metastatic BC model). The suppressive effect of miR-143-3p on the expression of this oncogene as a direct effect and PDL1 level as an indirect result in BC cell lines was investigated. Besides, the effect of miR-143-3p overexpression on BC cell lines proliferation through downregulation of candidate oncogene was investigated.

First, the targeting of miR-143-3P on MUC1 3 'UTR was confirmed by using bioinformatics software and then by dual luciferase assay. The expression level of miR-143-3P and MUC1 were measured in breast cancer lines compared to the normal line. After transfection of miR-143-3P into breast cancer cell lines, MUC1 and PD-L1 expression in mRNA levels were evaluated by using qRT-PCR. The miR-143-3p effects on MUC1 and PD-L1 expression and its impact on cell proliferation was investigated by using MTT assay, (mean ± SD).

The result of the dual luciferase assay was obtained from cloning the 3'UTR of the MUC1 gene in the psi-check2 vector and sequencing. According to the prediction analysis, the 3′-UTR of MUC1 gene has been considered as a putative miR-143-3p binding site (Target scan database). Transfecting miR-143-3p into MDA-MB-231, BT-549, MCF-7 cells harboring MUC1 3′-UTR decreased the luciferase activity to to 55% ± 1/73, 57% ± 1 /15 , 59 % ± 3/51% respectively, in comparison with the control (P < 0.01) (mean ± SD). he expression of miR-143-3p was significantly downregulated in BC cell lines compared to their control, and its downregulation was negatively correlated with MUC1 overexpression (r= -0.6419, p = 0.0244). Increased expression of miR-143-3P, decreased the mRNA level of MUC1 in MDA-MB231 to 1/3 ± 0/7, BT549 to 1/1± 0/45 and in MCF7 to 0/4/0 ±21, compared whit untransfected cells.
MiR-143-3p introduction to BC cells downregulates PDL-1 indirectly. Overexpression of miR-143-3P reduces the MUC1expression, and the reduction of MUC1 expression reduces PD-L1 expression mRNA level in MDA-MB231 to 3/4±0.45, BT549 to 3/1±0.56, MCF7 to 0/53±0.48, compared whit un transfected cells (fig4A). miR-143-3p inhibits BC cellular proliferation. An MTT assay revealed that miR-143-3p overexpression significantly suppressed the proliferation in MCF-7 to 52% ± 1.55, BT549 to 48% ± 2.45 MDA-MB231 to 43% ± 3.36, compared whit non-transfected cells 72 h after transfection.

 Breast cancer treatment has made significant progress in recent years, and a new treatment called immunotherapy, which uses the body's immune system to treat cancer, has become popular (6). One of the symptoms of breast cancer tumors is the overexpression of MUC1. A high expression level of MUC1 is associated with poor prognosis in cancer patients (7, 8). In an animal model of breast cancer, intravenous injection of MUC1 suppresses the immune system and accelerates the death of tumor-bearing mice. The serum level of MUCl expression is also associated with suppression of the immune system in patients with metastatic adenocarcinoma treated with active specific immunotherapy (8, 9). As we mentioned earlier, MUC1 can increase the expression of PD-L1 by recruiting MYC and NF -kB p65 to the PD-L1 promoter in some solid tumors, such as triple negative breast cancer (10). the mechanisms underlying PD-L1 expression in MUC1-positive tumor tissue were investigated, and the potential of targeting PD-L1 to inhibit the progression of MUC1-overexpressing colon cancer in mice, Therefore, the role of MUC1 in tumor immune evasion has been confirmed and can be a basis for using PD-L1 inhibitors in MUC1-positive colon cancer (8). microRNAs can regulate the expression of immune checkpoint receptors and their ligands (11). Changes in the expression of miR-143 often play a role in tumorigenesis (12). In 2020, Y Tokumaru and his colleagues showed by studying breast cancer samples with estrogen receptors that the high expression of microRNA-143 is associated with a favorable tumor immune microenvironment and better survival in breast cancer. They reported that miR-143 is a tumor suppressor microRNA that exhibits anti-tumor function by targeting KRAS signalling pathways in various malignancies. The high expression of miR-143 in cancer cells with a favorable tumor immune microenvironment regulates the positivity of anti-cancer immune cells. The suppression of pre-cancerous immune cells is related and leads to better survival of breast cancer patients (13).
In this study, we demonstrated MUC1 was potential as a novel target of miR-143-3p, therefore, the dual-luciferase reporter assay verified that miR-143-3p directly targeted MUC1 and inhibited this transcription. We found miR-143-3p downregulated in breast cancer cell lines compared to the normal line via qRT-PCR. In the current study, the qRT-PCR results illustrated an inverse relation between MUC1 mRNA expression and miR-143-3p expression in breast cancer cell lines. We investigated indirect targeting of PD-L1 via MUC1 targeting by miR-143-3p., be able to be prevent the cell proliferation in breast cancer cell line. Therefore we recognized, targeting MUC1 may be a novel strategy for BC therapy and prevent the progression of breast cancer and miR-143-3p as a promising candidate for miR restoration therapy in BC patients.

Recently, the use of immunotoxins for targeted cancer therapy has been proposed, to find new anticancer drugs with high efficacy on tumor cells with minimal side effects on normal cells. we designed and compared several arazyme (AraA)-based fusion proteins with different ligands to choose the best-targeted therapy for interleukin 13 receptor alpha 2 (IL13Rα2)-overexpressed cancer cells. For this purpose, IL13Rα2 was selected as a receptor and IL13 and IL13.E13K were evaluated as native and mutant ligands, respectively. In addition, Pep-1 and A2b11 were chosen as the peptide ligands for targeted cancer therapy.

Experimental approach: 

Several bioinformatics servers were used for designing constructs and optimization. The structures of the chimeric proteins were predicted and verified by I-TASSER, Q-Mean, ProSA, Ramachandran plot, and Verify3D program. Physicochemical properties, toxicity, and antigenicity were predicted by ProtParam, ToxinPred, and VaxiJen. HawkDock, LigPlot+ , and GROMACS software were used for docking and molecular dynamics simulation of the ligand-receptor interaction.

The in silico results showed AraA-A2b11 has higher values of confidence score and Q-mean score was obtained for high-resolution crystal structures. All chimeric proteins were stable, non-toxic, and non-antigenic. AraA-(A(EAAAK)4ALEA(EAAAK)4A)2-IL13 retained its natural structure and based on ligand-receptor docking and molecular dynamic analysis, the binding ability of AraA-(A(EAAAK)4ALEA(EAAAK)4A)2-IL13 to IL13Rα2 was sufficiently strong. 

Conclusion and implications:

 Based on the bioinformatics result AraA-(A(EAAAK)4ALEA(EAAAK)4A)2- IL13 was a stable fusion protein with two separate domains and high affinity with the IL13Rα2 receptor. Therefore, AraA-(A(EAAAK)4ALEA(EAAAK)4A)2-IL13 fusion protein could be a new potent candidate for target cancer therapy.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S) glycoprotein that projects from the virus surface is highly immunogenic. It is considered to be the target of many neutralizing antibodies as well as a target in vaccine design efforts. Evaluation the immunogenicity of a recombinant fragment of the spike protein (rfsp) that is comprised of Receptor Binding Domain (RBD), S1/S2 cleavage site, and fusion peptide (FP) as immunogenic proteins of SARS-COV-2, in BALB/c mice and evaluation of the efficacy of epitopes rfsp as a multi-subunit chimeric vaccine.

The present study made use of CHO-K1 (Chinese hamster ovary K1) cells to create a cell line for constant expression rfsp. The rfsp was purified with Ni-NTA chromatography and confirmed by Western blotting. The immunogenicity and neutralizing antibody efficacy of rfsp were evaluated in BALB/c mice. ELISA was employed to test rfsp via sera of COVID-19 convalescent patients infected with SARS-CoV-2 alpha and delta variants.

Our results showed significant differences in antibody titers in immunized mice compared to the control groups and neutralizing antibodies were positive, sera from mice immunized are capable of bound SARS-CoV-2 virus, chimer peptide is capable bound antibodies patients infected with SARS-CoV-2 and patients infected with delta variant SARS-CoV-2.

Overall, these results indicate that rfsp protein would be a novel potential antigen candidate for the development of a subunit SARS CoV-2 vaccine and rfsp has the potential to be a useful option for the development of the assays for serodiagnosis of SARS-CoV-2 infection.

Genome editing is an efficient and accurate method in biological and medical studies. Among the wide range of genome editing techniques, clustered regularly interspaced short palindromic repeats (CRISPR) is one of the simplest and yet promising methods. The CRISPR system consists of two key components; an endonuclease enzyme called Cas9 and guide RNA (gRNA), ensuring that Cas9 enzyme cuts at the right point in the genome. Although CRISPR genome editing is one of the useful methods to modify the genome, there are still multiple challenges in the technique steps.

sgRNAs were designed and ordered accordingly on the basis of the target site and vector of interest. Cloning procedures were performed and confirmed by sequencing as a gold standard. The list of high efficient sgRNAs for the LAMP-2 gene was prepared based on the available outstanding databases by analyzing and comparing the specificity and functionality as two main characteristics.

In this study, we tried to evaluate the main challenges in the process of preparing CRISPR/Cas9 popular vectors (pX330/pX459). A list of high efficient sgRNAs for an autophagy gene is also prepared as an example for clarification of the sgRNA design procedures.

This study tried to depict problems that may be encountered during sgRNA design and plasmid preparation, followed by giving appropriate recommendations.

The protective role of astaxanthin on human health, including the prevention of cardiovascular disease, prevention of cell aging, refers to the very strong antioxidant properties of this substance. Cadmium is one of the pollutants in ecosystems. This heavy metal acts as a toxin on the reproductive system and destroys the process of spermatogenesis in testicular tissue. The aim of this study was to evaluate the effect of astaxanthin on cadmium chloride induced testicular damage in adult male Wistar rats. In this study, 45 rats were randomly divided into 9 groups. The healthy control group did not receive any treatment. Control group, physiological serum, infertile control group, cadmium chloride (1 mg / kg body weight) by intraperitoneal injection, astaxanthin receiving groups (5, 10 and 20 mg / kg body weight) by gavage and groups They received cadmium chloride and astaxanthin (5, 10 and 20 mg / kg) at the same time. Test samples from testicular tissue homogenates were used to measure oxidative stress parameters. The parameters were analyzed by photometric method with ELISA reader and microplate reader. The results showed that astaxanthin (10 and 20 mg / kg body weight) increased the antioxidant enzymes SOD, GPX and CAT and astaxanthin (10 mg / kg body weight) decreased MDA. Thus, astaxanthin may act as an antioxidant in protecting the rat testis against oxidative stress induced by cadmium chloride.

Iran is one of the most important hot spots for cutaneous leishmaniasis (CL) in the world. To date, no studies have been done on both epidemiological aspects along with the length of the treatment course of CL. This study aimed to determine the relative frequency of CL in patients with suspected skin lesions and the duration of healing after treatment with different regimens.

This cross-sectional study was conducted on patients with CL referred to the skin diseases and leishmaniasis research center (SDLRC) in Isfahan during the years 2018 to 2019. Among 389 patients with suspected skin lesions, 150 cases were included with proven CL. Information such as age, sex, education, location, size of the lesion, duration of treatment, and the rate of recovery were recorded. SPSS software version 20 was used for data analysis, the chi-square, Fisher´s Exact, and one Way ANOVA tests were used with a significant level of p < 0.05.

Among 350 admitted cases, 150 cases were CL. positive (42.85%). The rate of complete recovery was higher in cases with an average age of 33.55 ±18.9 years, but these differences were not statistically significant (P =0.077). There was 34 cases more than the other groups in this range of age. ( The rate of complete recovery in patients with a history of migration to endemic areas was higher than in patients without a history of migration (P = 0.81)). The rate of complete recovery in patients whose means treatment duration was 59.03 ± 41.43 days was higher than other recovery periods (P = 0.23).

The rate of complete recovery was higher in adult cases than the other groups. In this study, it was proved that the rate of recovery of patients had the significant relationship with the average duration of treatment.

The severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), giving rise to the coronavirus disease 2019 (COVID-19), has become a danger to wellbeing worldwide. Thus, finding efficient and safe vaccines for COVID-19 is of great importance. As a basic step amid contamination, SARS-CoV-2 employs the receptor-binding domain (RBD) of the spike protein to lock in with the receptor angiotensin-converting enzyme 2 (ACE2) on host cells. SARS-CoV-2 receptor-binding domain (RBD) is the main human antibody target for developing vaccines and virus inhibitors, as well as neutralizing antibodies. A bacterial procedure was developed for the expression and purification of the SARS-CoV-2 spike protein receptor-binding domain.

In this research study, RBD was expressed by Escherichia coli and purified with Ni-NTA chromatography. Then it was affirmed by the western blot test. The immunogenicity and protective efficacy of RBD recombinant protein were assessed on BALB/c mice. Additionally, RBD recombinant protein was tested by ELISA utilizing sera of COVID-19 healing patients contaminated with SARS-CoV-2 wild type and Delta variation. 

Indirect ELISA was able to detect the protein RBD in serum of the immunized mouse expressed in E. coli. The inactive SARS-CoV2 was detected by antibodies within the serum of immunized mice. Serum antibodies from individuals recovered from Covid19 reacted to the expressed protein.

Our findings showed that RBD is of great importance in vaccine design and it can be used to develop recombinant vaccines through induction of antibodies against RBD.

Probiotic Lactobacillus spp. modulate immune response via interactions of their binding proteins with epithelial cells. We studied the presence of attachment protein-encoding genes (mub1, mub2, and mapA) in Lactobacillus strains with probiotic features isolated from inflammatory bowel disease (IBD) patients and their attachment strength relative to healthy individuals.

Bacterial strains have been isolated from stool samples of 35 healthy and 23 IBD volunteers. Lactobacillus spp. were identified using PCR. Strains with probiotic features were determined by testing resistance against acid and bile. Isolates were assigned as non-adhesive, adhesive, and strongly adhesive strains based on the number of attached bacteria to epithelial cells. Finally, PCR was used to detect the presence of mub1, mub2, and mapA genes.

Probiotic lactobacilli were isolated from 35/35 and 9/23 of healthy and IBD individuals and yielded a total of 87 and 28 strains, respectively. The Mub1 gene was detected in 95.4% and 100% (p>0.05), mub2 in 95.4% and 89.3% (p>0.05), and mapA in 94.3% and 78.6% (p<0.05) of healthy and IBD isolates, respectively. The numbers of bacteria attached to epithelial cells in healthy and IBD isolates were respectively 33.68±6.00 and 12.23±3.87 in non-adhesive, 71.3±10.83 and 42.17±1.33 in adhesive, 124.40±8.59 and 104.67±5.50 in the strongly adhesive group (p< 0.05).

Less Lactobacillus spp. with weaker attachments to epithelial cells colonize the gut in IBD than healthy individuals. These findings suggest the beneficial role of probiotics in the management of IBD.

To prevent pneumococcal infections, especially meningitis and bacteremia, and to overcome the serotype-dependent limitation of polysaccharide-based vaccines, the development of conserved protein-based vaccines is essential. This study aimed at investigate the in-silico analysis and epitope mapping of pneumococcal DnaJ for the first time, and to design the multi-epitope based vaccines with different categories by focusing on induction of both humoral and cellular immunities.

We predicted B- and T-cell epitopes, IL-4, IL-17, IL-10, and IFN-γ inducer epitopes of DnaJ using Immunoinformatics tools. The immunogenicity and conservation score of the predicted epitopes among pneumococcal prevalent clinical serotypes, the immune simulation of DnaJ administration in mammals and potential regions involved in DnaJ-TLRs interactions were analyzed. Finally, we proposed three classes of multi-epitope DnaJ-based vaccine candidates.

This protein had 24 and 15 predicted linear B-cell and helper T-cell epitopes, respectively, with a conservation score of 86-100% among prevalent clinical pneumococcal serotypes. DnaJ also had many IL-4 and IFN-γ inducing epitopes and was considered an IL-10 and IL-17 inducer protein. The immune simulation showed induction of both humoral and cellular immunity against DnaJ. The residues at positions 274, 280, 292, 297, 300, 316-319, 333, 336-340, 358,  363-366,  and 372 were predicted to be involved in DnaJ-TLR2 and DnaJ-TLR4 interactions. Three classes of proposed DnaJ-based constructs were based on only B-cell epitopes, only helper T-cell epitopes, and multi-epitopes of B- and T-cell and IL-17 epitopes.

The results showed that although DnaJ has been reported to play an important role in cellular immunity, our results indicated the high potential of DnaJ to stimulate mucosal, humoral, and cellular immunity.

Malignant breast cancer (BC) frequently contains a rare population of cells called cancer stem cells which underlie tumor relapse and metastasis, and targeting these cells may improve treatment options and outcomes for patients with BC. The aim of the present study was to determine the effect of silibinin on the self-renewal capacity, tumorgenicity, and metastatic potential of mammospheres.

The effect of silibinin on viability and proliferation of MCF-7, MDA-MB-231 mammospheres, and MDA-MB-468 cell aggregation was determined after 72-120 hours of treatment. Colony and sphere formation ability, and the expression of stemness, differentiation, and epithelial-mesenchymal-transition (EMT)-associated genes were assessed by reverse transcription-quantitative polymerase chain reaction (qRT-PCR) in mammospheres treated with an IC50 dose of silibinin. Additionally, the antitumor capacity of silibinin was assessed in vivo, in mice.

The results of the present study showed that silibinin decreased the viability of all mammospheres derived from MCF-7, MDA-MB-231, and MDA-MB-468 cell aggregation in a dose-dependent manner. Colony and sphere-forming ability, as well as the expression of genes associated with EMT were reduced in mammospheres treated with silibinin. Additionally, the expression of genes associated with stemness and metastasis was also decreased and the expression of genes associated with differentiation were increased. Intra-tumoral injection of 2 mg/kg silibinin decreased tumor volumes in mice by 2.8 fold.

The present study demonstrated that silibinin may have exerted its anti-tumor effects in BC by targeting the BC stem cells, reducing the tumorgenicity and metastasis. Therefore, silibinin may be a potential adjuvant for treatment of BC.

Akkermansia muciniphila, an important member of the gastrointestinal microbiota, is involved in the functioning of the host immune system. We aimed to investigate the effect of A. muciniphila and its OMVs on the expression of microRNA-21 and microRNA-34a involved in the maturation of human dendritic cells. A. muciniphila was cultured on a mucin-containing medium and its OMVs were extracted by ultracentrifugation. Extraction of peripheral blood mononuclear cells (PBMCs) was performed using Ficoll’s density gradient centrifugation method. Monocytes were differentiated into dendritic cells in the presence of interleukin-4 cytokines and granulocyte-monocyte colony-stimulating factor. Then, A. muciniphila at MOI 50 and 100, OMVs at a dose of 50 µg/µl, dexamethasone (to induce tolerogenic dendritic cells), and the LPS (to induce mature dendritic cells) were incubated. Finally, the expression of microRNA-21 and microRNA-34a was measured by real-time PCR. The morphological characteristics of the extracted OMVs showed that their size was between 20-200 nm. In LPS treatment, there were longer cytoplasmic growths than OMVs of dexamethasone and A. muciniphila groups. MicroRNA-21 expression was decreased in the LPS group compared to dexamethasone, A. muciniphila, and OMV groups, although this decrease was not significant (P> 0.05). In addition, microRNA-34a expression was significantly increased in the LPS group, which was reversed by treatment with dexamethasone, A. muciniphila, and OMV (P <0.001). MicroRNA-21 and microRNA-34a can exhibit anti-inflammatory and pro-inflammatory functions. A. muciniphila and its OMVs appear to be a viable option for introduction as a new generation probiotic.

Consuming a high-fat diet, as a stressor, during pregnancy and lactation can change the programming of the neuroendocrine system, and cause long -lasting adverse metabolic and behavioral effects in later life. In this study, the effect of high-fat diet feeding during pregnancy-lactation on basal plasma corticosterone level in adult male rat offspring was investigated. This study was performed in 1398, at Physiology Department, Shahid Beheshti University of Medical Sciences.

In this experimental study, dams received high-fat and normal diets from the beginning of pregnancy to the end of lactation. At the end of weaning, male offspring were divided into high-fat diet (HFD) and normal diet (ND) (6 rats per group according to previous studies) based on the dams’ diet and received normal diet until six weeks of age. Then the body weight of the animals was measured using a digital balance and blood samples were taken from their tails to determine the plasma concentration of corticosterone by a rat corticosterone Elisa Kit. Subsequently, their adrenal glands removed and weighed using a digital balance. The results were analyzed using independent t test.

Compared with the offspring of the ND group, the plasma concentration of corticosterone was higher in the offspring of the HFD group (2.11±0.13) (P<0.01), whereas the body weight of these offspring (114.8±2.37) (P<0.01) was lower. However, the adrenal gland weight of HFD group (28.25±2.80) was not significantly different from that of ND group (29.52±1.68). There were not any reduction in the samples of the study groups.

  Regarding the unchanged adrenal glands weight of the HFD group, it seems that the increase in plasma concentrations of corticosterone in this group is due to impaired corticosterone clearance.

Today, vitamin D deficiency (VDD) is one of the major health issues around the world and VDD is associated with several diseases. This study was conducted to find the relationship between vitamin D status in male’s serum with sperm function and clinical outcomes in infertile men candidate for intracytoplasmic sperm injection (ICSI).

In this cohort study, different parameters of male fertility such as sperm parameters, oxidative stress, and sperm chromatin status were evaluated in sperm samples of 30 infertile couples candidate for ICSI. Clinical outcomes like fertilization, embryo quality, and implantation were also assessed. Data were analyzed using SPSS Statistics 25.0 software. Besides, assessment of the correlation between aforementioned parameters with the level of serum vitamin D, in this study, ICSI candidates were divided into three groups [individuals with sufficient vitamin D levels (>30 ng/ml), insufficient vitamin D levels (between 20-29 ng/ml), and VDD (<20 ng/ml)]. The aforementioned parametesr were also compared between these study groups.

Analysis of all the data revealed a significant correlation between the level of vitamin D with sperm concentration (P=0.000, r=0.5), sperm count (P=0.03, r=0.31) and sperm reactive oxygen species (ROS) level (P=0.000, r=-0.77). Moreover, comparing clinical outcomes within study groups showed a significant difference in implantation rate between sufficient and other groups (insufficient and deficient) (P=0.02).

Considering the association between sperm concentration and level of ROS with vitamin D and, higher implantation rate in individuals with vitamin D sufficient group compared to other two groups, our data call for vitamin D supplementation as part of male infertility treatment. But considering our sample size, further research is needed to verify these findings.

The significant role of oxidative stress in the occurrence and development of a variety of diseases, including renal ischemia-reperfusion injury, has been thoroughly studied in this research. In this study, the protective role of indole-acetic acid on antioxidant, apoptotic and histopathological parameters in a rat model of renal ischemia-reperfusion (IR) injury were investigated.

We divided 40 rats into the following four groups (n = 10 per group): healthy control, IR control, IR + indole-acetic acid 40 mg/kg, and IR + indole-acetic acid 60 mg/kg. After two weeks, the rats were anesthetized and their kidneys were removed. The effects of indole-acetic acid on biochemical parameters [glutathione peroxidase (GPx) and catalase (CAT) were measured by spectrophotometry and expression of apoptotic genes (BAX and Bcl2) using real-time RT-PCR. Tubular necrosis was evaluated using a histopathological study.

There were significant improvements in biochemical parameters (GPx), expression of the apoptotic genes (BAX) and tubular necrosis in rats treated with indole-acetic acid.

Indole-acetic acid could reduce the effects of factors involved in the pathogenesis of IR, including oxidative stress, apoptosis and tubular necrosis. It can be recommended that, indoleacetic acid may be useful for amelioration of damages caused by IR.

Bacteriocins are antimicrobial peptides produced by many genera of bacteria especially Lactobacillus spp. against many pathogens, adapt bacterial composition in the gut and inhibit dysbiosis that can lead to inflammation disorders like inflammatory bowel disease (IBD). The aim of this study was to compare the prevalence of bacteriocin genes in health, IBD disease and recovery conditions.

In this survey 115 Lactobacillus spp. from 58 fecal samples of three different groups were evaluated. Comparison of the presence of bacteriocin genes in different groups were assayed by purified samples and PCR method, followed by statistical analysis to identify the effect of inflammation in the proportion of Lactobacillus spp. and presence of their bacteriocin genomes.

Of 115 Lactobacillus spp. 60% of samples had positive bacteriocin-encoding genes which included: gassericin-A 29.56%, acidocin 15.65%, plantaricin-NC8 18.26%, plantaricin-S 13.04%, lactacin-F 9.5%, sakacin-P 6.08% and gassericin-T 6.08%. Results indicated that the percentage of positive bacteriocin genes were much more in healthy volunteer and IBD-recovered in comparison to IBD-patients which showed the effect of inflammation in the presence of bacteriocin genes.

The results obtained in this study demonstrated that the presence of bacteriocin genes can be related to health and disease states and inflammatory disease affected the prevalence of bacteriocin-encoding genes. This approach can help to identify bacterial functions that can be targeted in future concepts of IBD therapy.

Multiple sclerosis (MS) is one of the most common diseases of the nervous system, characterized by inflammation of the central nervous system and oxidative stress. Carvacrol is a monoterpenoid phenol with antioxidant effects against free radical. The aim of this study was to evaluate the effect of carvacrol on the expression of Hmox-1, iNOS, Nrf2 and NF-ҚB genes in the spinal cord of Experimental Autoimmune Encephalomyelitis (EAE) rats as an animal model of MS.

EAE was induced in female Lewis rats and they were then divided into three groups: control, EAE model, and EAE treated with carvacrol. Carvacrol was daily injected intraperitoneally for 17 days after immunization. RNA was extracted from rat spinal cord tissue and changes in the expression of the Hmox-1, iNOS, Nrf2 and NF-ҚB genes were examined by Real-Time PCR.

The results showed that carvacrol treatment stopped severe weight loss in the EAE model group and their mean weight increased. Furthermore, carvacrol was found to reduce the expression of iNOS and NF-ҚB genes and increase the levels of Hmox-1 mRNA and Nrf2 mRNA.

The effect of carvacrol on weight changes in treated rats, along with the increased HO-1 gene expression in their spinal cord, associated with increased Nrf2 mRNA and decreased expression of iNOS and NF-ҚB, led to a reduction in inflammation and oxidative stress, and an increase in the neuroprotection. Furthermore, the invasive environment created for some cells, such as oligodendrocytes, was modulated.

PD-L1 is one of the most important immune control molecules in breast cancer and plays an important role in suppressing the immune system against tumor cells. Controlling the expression of PD-L1 at mRNA level using miRNA inhibitors could be helpful strategy for cancer treatment. In this study, considering the possible role of miR-145 as a tumor suppression in breast cancer, its involvement in reducing PD-L1 expression in breast cancer cell lines has been investigated.

First, the targeting of miRNA-145 on 3 'UTR of PD-L1 gene was confirmed using bioinformatics software and then by luciferase dual reporter assay. The expression level of miRNA-145 was measured in breast cancer cell lines compared to normal line. After transfection of miRNA-145 into breast cancer cell lines, qRT-PCR was used to evaluate the effect of miRNA-145 on PD-L1 expression.

we showed that decreased expression of miRNA-145 in breast cancer cell lines was directly related to increased PD-L1 expression (r= -0.6175, P value₌0.0457). In addition, increased expression of miRNA-145 in breast cancer cell lines MDA-MB231, BT549 and MCF7 significantly reduced PD-L1 expression (1.938±0.212, 1.784±0.03 and 0.083±0.02 respectively).

Our findings suggest that miRNA-145, by targeting the PD1/PD-L1 pathway and reducing PD-L1 expression, may be a therapeutic agent to prevent the progression of breast cancer.