ABSTRACT
Background: salivary gland tumors are among malignancies that have high recurrence rate. Immune responses in salivary gland tumors have not been well elucidated. T helper type 1 [Th1] and Th2 cytokines have been reported to play a role in the outcome of head and neck cancers
Objective: to evaluate the serum levels of interferon gamma [IFN- [gamma]], as the hallmark of Th1 cytokines, and interleukin-4 [IL-4], as the hallmark of Th2 cytokines, in patients with benign and malignant salivary gland tumors in comparison with healthy controls
Methods: fifty patients with benign and 14 patients with malignant salivary gland tumors, as well as 23 healthy individuals were recruited. Serum levels of IFN-[gamma] and IL-4 were measured using ELISA method. Nonparametric tests were used for data analysis
Results: serum levels of IFN-[gamma] and IL-4 were found not to be significantly different in patients compared to the control group [0.68 +/- 0.29 vs. 1.03 +/- 0.57 pg/ml, p=0.58 for IFN-[gamma], 4.57 +/- 1.57 vs. 4.41 +/- 1.31 pg/ml, p=0.28 for IL-4]. IFN-[gamma] and IL-4 serum levels were also not significantly different between patients with benign and malignant salivary gland tumors [p=0.54 and p=0.86, respectively]
Conclusion: the systemic levels of IL-4 and IFN-[gamma] seem not to be associated with salivary gland tumor in our study. Investigation of other cytokines produced by Th1 and Th2 cells are warranted
ABSTRACT
Evidences suggest that besides the neurotransmitters contributing to the development of depression, renin-angiotensin system [RAS] may also have a substantial role. Certain polymorphisms of RAS are associated with over activity of RAS and depression. Considering that antidepressants reduce the actions of angiotensin II, the main product of RAS, this may come into mind that genetic polymorphisms of the mentioned system may affect the outcome of therapy in depressed patients. In the present study, 100 newly diagnosed depressed patients, according to DSM-IV criteria, were treated with 20 mg of fluoxetine for 8-12 weeks. Patients were categorized into responsive and non-responsive groups according to 50% reduction in symptoms. Genotype frequencies of angiotensin-converting enzyme [ACE] gene [ACE [I/D, A-240T and A2350G]] were then determined in DNAs extracted from venous blood of the patients using polymerase chain reaction-restriction fragment length polymorphism [PCR- RFLP] and PCR. Results indicate that polymorphisms studied and their haplotypes were not associated with better response to fluoxetine. However, a strong association between age and treatment in depressed Iranian patients was observed [P=0.001]. In conclusion, unlike previous reports, this study does not support the hypothesis of special genotypes of RAS contributing to a better response to antidepressants in depressed patients
ABSTRACT
Klebsiella pneumoniae is a hospital-acquired pathogen that leads to various infections. Hence, efforts to develop an effective vaccine against that pathogen are well documented. Our interest is the production of the previously designed multi-epitope vaccine construct against the K. pneumoniae in a prokaryotic host. Therefore, a new construct containing the nucleotide sequence of the novel vaccine was successfully expressed in Escherichia coli and then purified by Ni-NTA spin column. The purified recombinant protein can be considered as potential vaccine candidate for wet-laboratory analysis aiming to fight K pneumoniae
ABSTRACT
Various fixation and permeabilization techniques have been developed for detection of intracellular antigens by flow cytometry; however, there are few studies using flow cytometry to detect the frequency of intracellular nucleic acids, particularly RNA. We tested six different permeabilization methods in order to gain access to a high quality method with minimal damage to intracellular components focusing on 18S rRNA in HeLa cells. HeLa cells were fixed in 2% paraformaldehyde. A variety of detergents and enzymes including saponin, TritonX-100, Tween-20, NP40, Proteinase K, and streptolysin O were used to optimize a protocol of permeabilization for the flow cytometric enumeration of intracellular 18S rRNA. Treated cells were subjected to standard protocol of flow cytometric in situ hybridization in the presence of FITC-labeled sense and antisense probes to detect 18S ribosomal RNAs. Samples were then analyzed on a FACSCalibur flow cytometer. To evaluate cell morphology, following hybridization the cells were fixed on glass slide, covered with DAPI, and evaluated on a fluorescent microscope with appropriate filter sets. In comparison with other methods, maximum cell frequency in percentage and fluorescent intensity [M1=2.1%, M2=97.9%] were obtained when the cells were treated with 0.2% Tween-20 and incubated for 30 min [p=0.001]. Our study indicated that the highest levels of mean fluorescence could be obtained when the cells were treated with Tween-20. However, it should be taken into consideration that for a successful flow cytometric result, other interfering factors such as hybridization conditions should also be optimized
Subject(s)
Flow Cytometry , HeLa CellsABSTRACT
Background: An association between lung cancer and chemokines has been advocated in the recent years. This study aims at investigating the association between lung cancer and 16C/A single nucleotide polymorphism [SNP] [rs. 4359426] in C-C motif chemokine 22 [CCL22] as well as C1014T SNP [rs. 2228428] in C-C chemokine receptor type 4 [CCR4], which serves as the receptor for CCL22
Methods: Genotyping was performed in 148 lung cancer patients and 148 normal controls using Polymerase Chain Reaction- Restriction-Fragment Length Polymorphism [PCR-RFLP]. The data were verified by direct automated sequencing
Results: Frequencies of CC, CA and AA genotypes of 16C/A SNP in CCL22 gene were 112 [75.7%], 33 [22.3%] and 3 [2.0%] in patients, and 119 [80.4%], 24 [16.2%] and 5 [3.4%] in controls respectively. No significant differences were observed in genotype frequencies at this position between cases and controls [P=0.34]. Moreover, there was no significant association between CCL22 polymorphism and types of lung cancer in patients. The distribution of CC, CT and TT genotypes of C1014T SNP in CCR4 gene, was 76 [51.4%], 60 [40.5%] and 12 [8.1%] in patients, and 80 [54.1%], 49 [33.1%] and 19 [12.8%] in controls respectively. No statistically significant differences were observed in genotypes frequencies of CCR4 gene between patients and controls [P=0.24]. The genotype inherited by patients observed not to be associated with the type of lung cancer [P>0.05]
Conclusion: Results reveal that CCL22 gene polymorphism at position 16C/A and CCR4 gene polymorphism at position C1014T, appear not to be associated with susceptibility to lung cancer
ABSTRACT
CD8+ cytotoxic T lymphocytes have been recently divided based on their cytokine expression profile. To evaluate the percentages of CD8+lymphocytes and their effector subsets including Tc1, Tc2 and Tc17 in the tumor draining lymph nodes [TDLNs] of patients with breast cancer. Single cell suspensions were obtained from TDLNs of 42 patients with breast cancer. Staining of the cell surface markers and intracellular cytokines was performed using appropriate fluorochrome-conjugated antibodies. The data was acquired on a four-color flow cytometer and was analyzed by CellQuestPro software package. The percentages of different CD8+ cell subtypes [Tc1, Tc2 and Tc17] were quantified in CD8+T lymphocytes. The comparison was made between LN+ versus LN- patients, as well as patients in different clinico-pathological status. The percentage of Tc1, Tc2 and Tc17 subsets were not significantly different between LN+ and LN- patients. Despite no difference in the percentages of Tc1 cells in LN+ patients with infiltrative ductal carcinoma [IDC], the mean expression of IFN-gamma by Tc1 cells decreased significantly in comparison to LN- patients. On the other hand, the percentages of Tc2 and Tc17 effector subsets were increased in advanced stages [p=0.018 and p=0.009, respectively]. As the first study to investigate various effector subtypes of CD8+ lymphocytes in TDLNs of patients with breast cancer, our data collectively suggests a positive association between IL-17- and IL-4-producing CD8+ T cell percentages [Tc2 and Tc17] in TDLNs with breast cancer progression. Although the number of Tc1 cells seems not to be affected by cancer progression, down-regulation of IFN-gamma by these cells seems to be associated with tumor metastasis to TDLNs. These findings may have implications in cancer immunotherapy based on CD8+ effector subsets.
ABSTRACT
Polycystic ovary syndrome [PCOS] has been suggested to be linked with autoimmune processes. Laparoscopic ovarian electrocauterization has the potency to stimulate more autoimmune reactions in PCOS patients. In the present study, we considered anti-nuclear antibodies [ANAs] as the hallmark of autoimmune reactions, and investigated the serum level of these antibodies in 35 patients with PCOS [21-38 years old] pre and one-month after electrocauterization, and in 35 fertile healthy women [25-35 years old] as the control group. Serum levels of ANAs, as well as ANA subtyping, were investigated using the Enzyme-Linked Immunosorbent Assay [ELISA]. While 3 out of the 35 patients [8.6%] were positive for ANAs before electrocauterization, none of the controls was positive. The number of ANA-positive cases increased following electrocauterization [3 out of 35 [8.6%] before vs. 10 out of 35 [28.6%] after the procedure]. The main ANA subtype in the positive samples was SS-A. The higher ANA level among the PCOS patients suggests association of the disease with autoimmune reactions. Laparoscopic ovarian electrocauterization seems to increase the number of positiveANA patients
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A possible mechanism by which hyperthermia enhances tumor immunogenicity is the induction of NKG2D ligands on tumor cells. Although the expression of MHC class I chain-related protein A and B [MICA/B] has previously been reported in different carcinomas, there is no information about MICA/B expression in liposarcomas. To investigate MICA/B induction in a human liposarcoma cell line [SW-872] after thermotherapy. SW-872 and HeLa cell lines were subjected to thermal stress for 1 h at 42, 44 and 46[degree sign]C, and after 2, 4 and 6 h of incubation at 37[degree sign]C, MICA/B expression was assessed at the mRNA and protein levels. Despite high levels of MICA/B transcripts in SW-872 cells at baseline, the expression of these genes decreased significantly at both the mRNA and protein levels after almost all thermal treatments. Our data conclude that thermotherapy under 42-46[degree sign]C had no effect on MICA/B induction on SW-872 liposarcoma cell line but the effects of fever-range temperatures remain to be tested on this cell line
Subject(s)
Hyperthermia, Induced , NK Cell Lectin-Like Receptor Subfamily K/metabolism , Killer Cells, Natural/immunology , Cell Line, Tumor , Killer Cells, NaturalABSTRACT
T helper 1 and T helper 17 cells play important roles in immunity against foreign invaders. Differentiation of these Th subsets is affected by state of maturation and cytokines that are produced by dendritic cells [DCs]. Curdlan is a linear [1-3]-beta- glucan and has shown activity against tumors and infectious agents. This study aims to investigate whether curdlan plays its role through affecting the maturation and cytokine production by DCs. DCs were isolated from the spleen of BALB/c mice by MACS method. After an overnight culture of DCs in the presence of curdlan, the expression levels of CD40, CD86, and MHC-II molecules were determined by flow cytometry. The production of cytokines involved in Th1 and Th17 cell differentiation [IL-12 and IL-6, respectively] was also evaluated by ELISA. Lipopolysaccharide [LPS] treated and untreated cells were considered as positive and negative controls, respectively. The results of this study did not show a significant difference in the levels of surface expression of CD40 [p=0.82], CD86 [p=0.79], and MHC class II [p=0.84] molecules upon exposure to curdlan. However, LPS increased the intensity of CD40 expression on dendritic cells [p=0.04]. In addition, it was revealed that curdlan-exposed DCs are not able to produce a significant amount of IL-6 and IL-12 cytokines. Conversely, LPS-treated DCs were able to make a significant amount of IL-12 [p=0.005]. The results of the present study suggest that curdlan has no effect on Th1 or Th17 differentiation while LPS may induce Th1 deviation by induction of CD40 expression and IL-12 production.
ABSTRACT
Variations in Cytotoxic T Lymphocyte Antigen-4 [CTLA-4] affect the expression and function of this protein. We aimed to investigate the association of +49 A/G [rs231775], +1822 C/T [rs231779] and +6230 A/G [CT60, rs3087243] genetic variations, as well as the merged haplotypes in CTLA-4 gene with susceptibility to, or progression of head and neck cancer. Eighty patients with confirmed head and neck [HN] cancer [age 54.9 +/- 16.1 years] and 85 healthy age/sexmatched controls [age 56.3 +/- 12.4 years] were enrolled in the study. Genotypes were investigated by the PCR-RFLP method. Arlequin software package was used to check for Hardy-Weinberg equilibration, and to estimate the haplotypes. At position +6230 A/G [CT60], AA genotype, as well as A allele was significantly decreased in patients with HN cancers than controls [18.8% vs. 40.7%, p=0.004; odds ratio=0.34, and 46.3% vs. 61.7, p=0.007; odds ratio=0.53%, respectively]. Nearly the same results were obtained when we compared the subgroup of patients with squamous cell carcinoma of the HN [SCC-HN] with control subjects. The frequencies of genotypes and alleles at other positions were not significantly different between patients and controls, however ACG, GTA and GCA haplotypes emerged from three investigated loci occurred with significantly more frequencies in patients [p<0.0001], while ACA and GTG haplotypes were more frequent among controls [p<0.0001]. Significant differences of haplotypes, genotypes and alleles frequencies resisted the Bonferroni correction. Our results suggest that CT60 A allele, as well as ACA and GTG haplotypes in CTLA-4 gene may have protective roles against HN cancer in Iranian population, while ACG, GTA and specially GCA haplotypes may render susceptibility.
ABSTRACT
CCL22/MDC is a CC chemokine with a critical role in regulation of the immune balance in physiological condition. CCL22/CCR-4 ligation has been documented to participate in the migration of regulatory T [Treg] cells and Th2 lymphocytes to the site of breast tumors; circumstances that are known to be associated with poor prognosis. To investigate the association of a single nucleotide polymorphism [SNP] in CCL22 gene; 16C/A [rs4359426; Asp2Ala], with susceptibility to breast cancer in a sample of Iranian population. Methods: 161 patients with pathologically confirmed breast carcinoma [mean age 49.3 +/- 11.5 yrs] and 178 agematched healthy women [mean age: 49.3 +/- 12.9 yrs] were studied. CCL22 genotypes were investigated by the Polymerase Chain Reaction-Restriction Fragment Length Polymorphism [PCR-RFLP] method. Data was verified by direct automated sequencing. Arlequin analysis showed no deviation from Hardy-Weinberg equilibrium. The most frequent genotype in both patient and control groups was wild type CC genotype with frequency of 146 out of 161 [90.7%] among patients and 153 out of 178 [86.0%] in control group [p=0.24]. The frequency of CA genotype was 15 [9.3%] and 23 [12.9%] in patients and controls, respectively [p=0.38]. No AA genotype was observed among patients but this genotype was observed with the frequency of 2 out of 178 [1.1%] in control subjects. The minor allele frequency [MAF] was 0.07 in the population. No correlation was found between the investigated genotypes and clinicopathological characteristics of the patients. Conclusively, results of this investigation do not support the association of 16C/A SNP [rs4359426; Asp2Ala] in CCL22 gene with susceptibility to, and progression of, breast cancer in Iranian population
ABSTRACT
Myasthenia gravis [MG] is the most common disorder of neuromuscular junction in which autoantibodies develop against nicotinic acetylcholine receptor for unknown reasons. The association of immunomodulator genes with different autoimmune disease has been studied in recent years. The aim of this study was to investigate correlation between a genetic variation in Stromal Cell Derived Factor-1 [SDF1] and susceptibility to MG in an Iranian population. Genotyping of SDF1 at position 801 G/A was performed by Polymerase Chain Reaction-Restriction Length Polymorphism [PCRRFLP] in 87 patients with confirmed myasthenia gravis and 261 normal control subjects. No statistically significant differences were observed in the frequencies of genotypes and alleles between patients and controls [p>0.05]. Furthermore, no significant differences in the genotype distribution were found between the cases with different stages [p>0.05]. Our data suggest that the SDF1 gene polymorphism at position 801 G/A is not associated with myasthenia gravis
ABSTRACT
Ovarian cancer is a relatively common cancer among postmenopausal women. Nowadays, there is controversy about immunotherapy of ovarian cancer patients with interleukins such as interferon to reach better out come in prognosis of patients under chemotherapy. CTLA-4 is a gene, which has an important role in homeostasis and regulation of immune response. Inhibitory nature of CTLA-4 is proved to be of significance in autoimmune diseases as well as in cancer. In this study we intend to find out the relationship between polymorphisms of this gene at the sites of +49 A/G and -318 C/T and ovarian cancer. The polymorphisms of the CTLA-4 gene at the sites of +49 A/G exon and -318 C/T promoter were investigated. Blood samples of 73 patients with ovarian cancer and 115 healthy subjects used for DNA extraction. Two groups genotypes and alleles were determined using PCR method and compared by statistical t-student test. There was no statistically significant difference in genotypes and alleles prevalence of +49 A/G and -317 C/T between two groups [p>0.05]. Further researches with larger sample size while paying attention to the relation between the gene polymorphism and stage and type of tumor is recommended