Induction of apoptosis in cancer cells of pre-B All patients after exposure to platelets, platelet-derived microparticles and soluble CD40 ligand

Objective: The in vitro treatment of tumor cells with platelet [Plt] causes inhibition of tumor cell growth, although mechanism of this effect is not clear yet. Induction of apoptosis has been proposed as a mechanism of Plt effects on tumor cells. The purpose of this study was to clarify the role of Plts and Plt-derived components in the induction of apoptosis in the blood mononuclear cells of patients with leukemia

Materials and Methods: In this experimental study, peripheral blood mononuclear cells [PBMCs] were isolated from whole blood of five patients with childhood B-precursor acute lymphoblastic leukemia [pre-B ALL] and encountered with Plts, Plt-derived microparticles [Plt-MPs] as well as purified soluble CD40L [sCD40L]. After 48 hours of co-culture, the anti-cancer activity of the aforementioned factors was surveyed using examination of apoptosis markers of the cells including active caspase-3 and CD95 using ELISA and flow cytometer techniques, respectively. Additionally, staining of the cells with 7-Aminoactinomycin D [7-AAD] was evaluated by flow cytometer technique. Trypan blue exclusion test and WST-1 method were also used to compare the death/survival status of the cells

Results: Levels of CD95 and caspase-3 were significantly increased in the all treated groups [P<0.05]. On the other hand, trypan blue, 7-AAD and WST-1 methods showed significantly lower number of the live cells in the treated groups [P<0.05]

Conclusion: This study can show the ability of Plts, Plt-MPs and sCD40L for the induction of apoptosis in PBMCs of pre-B-ALL patients. Further studies are necessary to elucidate the different effects of platelets on cancer cells in vitro and in vivo

effect of cell derived microparticles in transfusion medicine and adaptive immune system

This article reviews will focus on the concept and formation of micro particles [MPs] in circulation and their role in transfusion medicine and immune system. MPs are cell membrane derived vesicles which express markers of their parent cells and are found in circulation at low levels. Exact functions of MPs are unclear. In here, Physiological almost all types of circulating MPs including platelets MPs [PMPs], leukocytes MPs [LMPs], red blood cells MPs [RMPs] and endothelial cells MPs [EMPs] have been discussed. Furthermore, MPs present in plasma and blood products and their levels increase during storage. Thus, it can be stated that MPs are likely to cause transfusion reactions, particularly thrombotic complications and Transfusion-Related Acute Lung Injury [TRALI]. Also, it is shown that the MPs may affect the immune system. However, to prove these, more and extensive studies both in vivo and in vitro need to be done

Conjugates of B-cells and platelet microparticles in the human peripheral blood

Background: Activated platelets shed microparticles [MPs] in vivo and certainly in vitro under storage. Like platelets, platelet-derived MPs contribute to hemostatic and inflammatory responses. We sought to determine the interactions between platelet MPs and peripheral B lymphocytes in the healthy blood circulation to propose a possible role for platelet MPs in the functioning of B cells

Materials and Methods: An enzyme-linked immunosorbent assay [ELISA] was established to determine the normal interactions between human peripheral blood B lymphocytes and platelet MPs. B cells were isolated and bound to the wells of microtiter plates using coated anti-CD19. Then the presence of attached MPs was surveyed. Also, platelet MPs were separated from human platelet concentrates and applied to confirm the new binding capacities of B cells for these microvesicles

Results: Platelet MPs were recognized in the wells of ELISA in which only B cells were isolated. So MPs were bound with peripheral blood B cells. Furthermore, using this method, the role of CD40/ CD40L interaction was displayed for the binding

Conclusion: It seemed that the binding of platelet MPs to B cells normally took place in vivo and a percent of B cells circulate in blood in connection with platelet MPs

Postoperative pain, nausea and vomiting among pre- and postmenopausal women undergoing cystocele and rectocele repair surgery / 대한마취과학회지

BACKGROUND: Postoperative nausea and vomiting (PONV) and postoperative pain are among the most common side-effects of surgery. Many factors, such as a change in the level of sex hormones, are reported to affect these complications. This study aimed to evaluate the probable effects of the menopause on PONV and postoperative pain. METHODS: Prospective study, in which a total number of 144 female patients undergoing cystocele or rectocele repair surgery under standardized spinal anesthesia were included. Patients were divided into two equally sized sample groups of pre- and postmenopausal women (n = 72). The occurrence of PONV, the severity of pain as assessed by visual analog scale (VAS) pain score, and the quantity of morphine and metoclopramide required were recorded at 2, 4, 6, 12, 18 and 24 h after surgery. RESULTS: The mean VAS pain score and the mean quantity of morphine required was higher among premenopausal women (P = 0.006). Moreover, these patients required more morphine for their pain management during the first 24 h after surgery compared to postmenopausal women (P < 0.0001). No difference was observed between the two groups regarding the incidence of PONV (P = 0.09 and P = 1.00 for nausea and vomiting, respectively) and the mean amount of metoclopramide required (P = 0.38). CONCLUSIONS: Premenopausal women are more likely to suffer from postoperative pain after cystocele and rectocele repair surgery. Further studies regarding the measurement of hormonal changes among surgical patients in both pre- and postmenopausal women are recommended to evaluate the effects on PONV and postoperative pain.

immunosuppressive activity of amniotic membrane mesenchymal stem cells on T lymphocytes

Mesenchymal Stem Cells [MSCs] are isolated from different sources like placenta. The placenta and its membranes like Amniotic Membrane [AM] are readily available and easy to work with. There is only limited knowledge on the immunomodulatory properties of human Amniotic Membrane-derived Mesenchymal Stem Cells [hAM-MSCs]. The aim of this study was to survey the suppressive activity of hAM-MSCs on T lymphocytes in vitro. Human AMs were obtained after caesarean section births from healthy women. After enzymatic digestion, cells were cultured and hAM-MSCs were obtained. In addition, human T lymphocytes were isolated and co-cultured with hAM-MSCs for 72 hr in the presence or absence of phytohemagglutinin [PHA]. Subsequently, proliferation of T cells was analyzed using BrdU and subsequently flow cytometry technique. Besides, the production of IL-4 and IFN-gamma was examined by ELISA method. Additionally, the expression of activation markers [CD38, HLA-DR] was studied on T lymphocytes by flow cytometry technique. It was revealed that hAM-MSCs could significantly suppress the proliferation of T lymphocytes [p

effect of sexual health education program on women sexual function in Iran

Sexual dysfunction is the most common disorder in women. According to the WHO, sexual education programs are considered as a need. Therefore, this study was designed to investigate the effect of educational program on sexual function in women with sexual dysfunction. This randomized trial, was conducted in 2013 on 90 married women by convenient sampling in Qazvin, central Iran. The demographic, Female Sexual Function Index [FSFI], and Beck's Depression Inventory [BDI] questionnaires were completed during structured interviews. After completing the sample size, subjects were divided randomly into two groups by using the table of random numbers [educational and control groups], then respectively received an educational intervention in the four sessions with one week interval and routine program offered by the center and following-up was done with refilling questionnaires 8 weeks after intervention. Sexual function improved after sex educational programs in all dimensions [sexual desire [P=0.006], sexual exciting [P=0.006], vaginal moisture [P=0.002], sexual satisfaction [P=0.011], and total score of sexual function [P=0.001]. Considering the importance role of sexual function in family strength, health, and development, it can be claimed that educational sex programs can help practitioners to improve sexual function of married women with sexual dysfunction

The caspase-3 inhibitor (peptide Z-DEVD-FMK) affects the survival and function of platelets in platelet concentrate during storage

BACKGROUND: Although apoptosis occurs in nucleated cells, studies show that this event also occurs in some anucleated cells such as platelets. During storage of platelets, the viability of platelets decreased, storage lesions were observed, and cells underwent apoptosis. We investigated the effects of caspase-3 inhibitor on the survival and function of platelets after different periods of storage. METHODS: Platelet concentrates were obtained from the Iranian Blood Transfusion Organization in plastic blood bags. Caspase-3 inhibitor (Z-DEVD-FMK) was added to the bags. These bags along with control bags to which no inhibitor was added were stored in a shaking incubator at 22degrees C for 7 days. The effects of Z-DEVD-FMK on the functionality of platelets were analyzed by assessing their ability to bind to von Willebrand factor (vWF) and to aggregate in the presence of arachidonic acid and ristocetin. Cell survival was surveyed by MTT assay. RESULTS: At day 4 of storage, ristocetin-induced platelet aggregation was significantly higher in the inhibitor-treated (test) than in control samples; the difference was not significant at day 7. There was no significant difference in arachidonic acid-induced platelet aggregation between test and control samples. However, at day 7 of storage, the binding of platelets to vWF was significantly higher in test than in control samples. The MTT assay revealed significantly higher viability in test than in control samples at both days of study. CONCLUSION: Treatment of platelets with caspase-3 inhibitor could increase their functionality and survival.

Effects of platelet microparticles on the activation of B cells

Platelets are anucleated fragments derived from megakaryocytes. It has been demonstrated that platelets play a role in hemostasis and innate immunity. In addition, platelets have a CD40 ligand which is an important molecular marker in motivating immune cells. Thus, platelets also have a role in adaptive immunity as seen by their ability to activate B cells. Since human platelet microparticles [MPs] originate from platelets, we have chosen to examine the effects of MPs on B cell activation. Platelet MPs were isolated from platelet concentrates obtained from the Tehran Blood Transfusion Center. The MPs were co-cultured with B cells isolated from human whole blood with magnetic beads using negative selection. After seven days, the expression of activation markers CD27 and CD86, as well as IgD were evaluated by flow cytometry. In a comparison between test [B cells/MPs] and control [B cells] cells we observed that the expression of activation markers CD27 and CD86 increased during the seven-day co-culture period. However, the expression of IgD antibody decreased. As with platelets, MPs can affect B cell activation during in vitro co-culture

Sequence analysis of the polymerase gene in hepatitis B virus infected blood donors in Iran

Hepatitis B is one of the most common infectious diseases worldwide that can be transmitted by blood transfusion. The hepatitis B virus [HBV] has eight different genotypes that show different geographical distributions and clinical manifestations. This study aims to investigate the sequence of the HBV polymerase gene and the frequency of HBV genotypes among Iranian blood donors. The sera of 223 blood donors who were positive for hepatitis B surface antigen [HbsAg] as determined by the ELISA method were selected. HBV DNA was extracted from the sera of 134 blood donors by a commercial kit, and the entire polymerase gene was amplified by nested-PCR. HBV genotypes were determined by direct sequencing of the HBV polymerase gene. Phylogenetic trees were constructed by the neighbor-joining [NJ] method. No known base mutations were found in the entire HBV polymerase gene of infected blood donors, and only genotype D was detected among HBV-infected blood donors. The sub-genotype D1 of HBV was dominant in the subjects. This study shows that antiviral-resistant mutations, such as lamivudine-resistant HBV strains, do not exist naturally among Iranian blood donors. More studies on the full-length HBV genomes are required to determine genome evolution of HBV among infected Iranian blood donors

[ relationship between soluble serum HLA-G and ICSI success rates]

Pregnancy is a successful transplantation. Factors evading rejection of the fetus by the mother's immune system are poorly understood and success rate and maintenance of embryos in assisted reproductive technologies [ART] may also depend on the same factors. The molecules of HLA-G are non-classical major histocompatibility complex class I antigens that have recently attracted attention in regards to pregnancy. The aim of the present study was to determine the concentration of HLA-G and its correlation with success or failure rates of ICSI. Serum samples of 107 women who were undergoing ICSI [the case group] were collected before and 14 days after embryo transfer, as were serum samples of 24 women with normal pregnancy [the control group] in the first trimester of pregnancy. Soluble HLA-G1 and G5 isoforms and the total sHLA-G were assayed by sandwich ELISA. Nonparametric Kolmogorov-Smirnov [K-S], Mann Whitney U and Wilcoxon tests were used for statistical analysis. No significant differences were observed in clinical variables including age, infertility duration and treatment regimen between the control and the case groups. Levels of sHLA-G1 and sHLA-G5 and the total sHLA-G prior and after ICSI in the control group, respectively, were 47.4 +/- 62.8 U/ml, OD: 1.47 +/- 0.58 prior and 59.6 +/- 69.5 U/ml, OD: 1.38 +/- 0.57 after ICSI. In the non-pregnant group, the values respectively were 35.7 +/- 55.2 U/ml, OD: 1.37 +/- 0.45 prior and 39.7 +/- 57.2 U/ml, OD: 1.31 +/- 0.46 after ICSI, corresponding to the control group; 53.16 +/- 47.92 U/ml and OD: 1.29 +/- 0.49. No significant statistical differences were found between the pregnant, nonpregnant and the control groups. No significant changes in the serum levels of sHLAG1 and sHLA-G5 isoforms and the total sHLA-G were observed following embryo transfer. No significant correlation was found between sHLA-G and the success of pregnancy in women undergoing ART. It seems that serum HLA-G has no prognostic value in the prediction of ICSI failure

[ preparation of albumin as a biological drug from human plasma by fiber filtration]

In recent years, consumption of whole-blood for the treatment of patients has decreased but use of biological plasma-derived medicines such as albumin, immunoglobulin and coagulation factors have increased instead. Paying attention to albumin molecular structure is important for its isolation from human plasma. Albumin is a single-chain protein consisting of about 585 amino acids and a molecular weight of 66500 Daltons. Albumin is a stable molecule and it is spherical in shape. There are different methods for human albumin preparation. Considering the large consumption of this biological drug in clinical settings, methods with fewer steps in production line are of big advantage in saving time and manufacturing more products. In this project, we prepared human albumin using hollow fiber cartridges in order to omit the rework on fraction V+VI. Human albumin is usually produced by the application of cold ethanol method, where albumin is obtained from fraction V by doing a rework on fraction V+VI to separate fraction V. In the current work, human albumin was prepared from fraction V+VI by the help of hollow fiber cartridges. With a concentration of 20%, the obtained albumin had 96.5% of monomer and 3.5% of polymer and polymer aggregate. Comparing the obtained human albumin with a number of commercial human albumin samples by the use of SDS-page, the results were satisfactory regarding the 3.5 percent polymer and aggregate rate for the prepared albumin

[Analysis of HLA-G transcripts in PBMCs of normal and lupus individuals and effect of IFN-gamma on the expression of HLA-G]

HLA-G is a nonclassical major histocompatibility complex antigen and is expressed as seven isoforms, including four membrane bound [HLA-G1 to-G4] and three soluble [HLA-G5 to-G7] forms. The pattern of selective expression of HLA-G transcripts in tissues shows the existence of a tight transcriptional control on the gene expression. It has been revealed that cytokines including interfrons and IL-10 could cause stimulation of the HLA-G transcription. The purpose of this study was to examine the effects of IFN-gamma on the expression of HLA-G transcripts in both PBMCs of normal and SLE patients. Whole blood of 20 female SLE patients and 15 healthy donor candidates for Bone Marrow Transplantation were used. PBMCs were isolated from the whole blood by Ficoll gradient centrifugation and cultured with or without IFN-gamma /LPS for 48 hours. Total RNA was extracted from the cells by trizol method. After reverse transcription of RNA to cDNA and the performance of a multiplex PCR for beta actin and HLA-G, the PCR products were analyzed using electrophoresis on a 2% agarose gel and stained with ethidium bromide. The results showed that the transcription of HLA-G was higher in SLE patients compared to normal controls. Addition of IFN-gamma /LPS could influence the expression of this molecule by increasing the transcription of HLA-G in both normal and patient PBMCs [P?0.05]. Transcription of HLA-G gene could be increased by the cytokine IFN-gamma. This observation is in accordance with previous reports. This effect could be assigned to both normal and lupus Patients. The effects of the cytokine IFN-gamma /LPS in the induction of HLA-G transcription were higher in normal than lupus patients. Nevertheless, the total expression of HLA-G was higher in lupus patients

Study on the frequent expression of rodgers and chido, red blood cell antigens, in iranian healthy population

Rodgers [Rg] and Chido [Ch] are blood-group antigens and they determine the fourthcomponent of human complement C4. Rodgers and Chido are associated with two C4 isotypes [C4A and C4B]. In addition to genotype determination, study on expression of Rg and Chido could be useful in disease studies. DNA was extracted from the whole blood of 60 normal individuals. Then, PCR amplification of C4d gene fragment was followed by restriction digestion. This study demonstrated that the frequency of Ch and Rg in Iranian healthy population was 98.3 and 93.4 percent, respectively. Additionally, 6.6 percent of the studied population showed Chido-positive, Rodger-negative and 1.7 percent showed Rodger-positive, Chido-negative genotype. It may be concluded that upon receiving blood transfusion, 6.6 and 1.7 percent of individuals could produce anti-Rg and anti-Ch antibodies, respectively

Null allele frequencies at HLA-G locus in Iranian healthy subjects

HLA-G gene contains 15 alleles including a null allele, HLA-G*0105N. Previous studies have shown that HLA-G*0105N does not encode the complete HLA-G1 or HLA-G5 isoforms but encodes a functional HLA-G protein with the ability to inhibit NK cell cytolysis. Thus, although the biological functions of HLA-G1 and HLA-G5 proteins are abrogated, other isoforms such as HLA-G2 can replace their roles. Studies on the null allele of HLA-G gene could be useful in understanding the genetic variants of HLA-G alleles in ethnic groups. The goal of this research was to determine the frequency of HLA-G*0105N null allele in Iranian healthy subjects. The frequency of HLA-G*0105N null allele was evaluated in Iranian healthy subjects by PCR-RFLP method. Genomic DNA was isolated from the whole blood of 100 randomly selected, healthy, unrelated Iranian individuals using salting-out technique followed by PCR amplification of the exon 3 of HLA-G gene. PCR products were digested with PpUM-1 and the resulted fragments were analyzed using gel electrophoresis. In this study the restriction enzyme digestion confirmed homozygous HLA-G*0105N null allele for 9% of the population. Furthermore obtained results indicated that the total frequency of HLA-G*0105N null allele was 20% in the studied population of Iran. The final data analysis showed that the total frequency of this allele in Iranian people was higher than other ethnic groups that have been studied so far

Effects of protoplast fusion on S-endutoxin production in bacillus thuringiensis Spp. [H14]

In this study, mutant forms of Bacillus thuringiensis spp. israelensis [H14] were produced. These mutants were identified when the cells were cultured on chloramphenicol plates and stained with crystal violet. Protoplasts of the mutants were isolated by enzymatic digestion [lysozyme] of the cell walls at the presence of an osmotic stabilizer. The protoplasts were induced to fuse to each other in the presence of PEG 6000. The frequency of regeneration and recombination was 80% and 2 10-4, respectively. In order to survey the effect of protoplast fusion on production of toxin, anti-serum against pure toxin was raised in rabbit and was used in single radial immunodiffusion. The comparison of -endotoxin concentration between B. thuringiensis fusion and the wild type strains showed that B. thuringiensis fusion has 1.48 time more toxin than wild type