Human Chorionic Villous Mesenchymal Stem Cells Modify the Functions of Human Dendritic Cells, and Induce an Anti-Inflammatory Phenotype in CD1+ Dendritic Cells.

BACKGROUND: Mesenchymal stem cells derived from the chorionic villi of human term placenta (pMSCs) have drawn considerable interest because of their multipotent differentiation potential and their immunomodulatory capacity. These properties are the foundation for their clinical application in the fields of stem cell transplantation and regenerative medicine. Previously, we showed that pMSCs induce an anti-inflammatory phenotype in human macrophages. In this study, we determined whether pMSCs modify the differentiation and maturation of human monocytes into dendritic cells (DCs). The consequences on dendritic function and on T cell proliferation were also investigated. METHODS: Interleukin-4 (IL-4) and granulocyte-macrophage colony stimulating factor (GM-CSF) were used to stimulate the differentiation of monocytes into immature dendritic cells (iDCs), which were subsequently co-cultured with pMSCs. Lipopolysaccharide (LPS) was used to induce maturation of iDCs into mature dendritic cells (mDCs). Flow cytometry and enzyme-linked immunosorbent assays (ELISA) were used to quantify the effect pMSC co-culturing on DC differentiation using CD1a, a distinctive marker of DCs, as well as other molecules important in the immune functions of DCs. The phagocytic activity of iDCs co-cultured with pMSCs, and the effects of iDCs and mDC stimulation on T cell proliferation, were also investigated. RESULTS: Monocyte differentiation into iDCs was inhibited when co-cultured with pMSCs and maturation of iDCs by LPS treatment was also prevented in the presence of pMSCs as demonstrated by reduced expression of CD1a and CD83, respectively. The inhibitory effect of pMSCs on iDC differentiation was dose dependent. In addition, pMSC co-culture with iDCs and mDCs resulted in both phenotypic and functional changes as shown by reduced expression of costimulatory molecules (CD40, CD80, CD83 and CD86) and reduced capacity to stimulate CD4(+) T cell proliferation. In addition, pMSC co-culture increased the surface expression of major histocompatibility complex (MHC-II) molecules on iDCs but decreased MHC-II expression on mDCs. Moreover, pMSC co-culture with iDCs or mDCs increased the expression of immunosuppressive molecules [B7H3, B7H4, CD273, CD274 and indoleamine-pyrrole 2,3-dioxygenase (IDO). Additionally, the secretion of IL-12 and IL-23 by iDCs and mDCs co-cultured with pMSCs was decreased. Furthermore, pMSC co-culture with mDCs decreased the secretion of IL-12 and INF-γ whilst increasing the secretion of IL-10 in a T cell proliferation experiment. Finally, pMSC co-culture with iDCs induced the phagocytic activity of iDCs. CONCLUSIONS: We have shown that pMSCs have an inhibitory effect on the differentiation, maturation and function of DCs, as well as on the proliferation of T cells, suggesting that pMSCs can control the immune responses at multiple levels.

Human placental mesenchymal stem cells (pMSCs) play a role as immune suppressive cells by shifting macrophage differentiation from inflammatory M1 to anti-inflammatory M2 macrophages.

BACKGROUND: Mesenchymal stem cells (MSCs) have a therapeutic potential in tissue repair because of capacity for multipotent differentiation and their ability to modulate the immune response. In this study, we examined the ability of human placental MSCs (pMSCs) to modify the differentiation of human monocytes into macrophages and assessed the influence of pMSCs on important macrophage functions. METHODS: We used GM-CSF to stimulate the differentiation of monocytes into the M1 macrophage pathway and then co-cultured these cells with pMSCs in the early stages of macrophage differentiation. We then evaluated the effect on differentiation by microscopic examination and by quantification of molecules important in the differentiation and immune functions of macrophages using flow cytometry and ELISA. The mechanism by which pMSCs could mediate their effects on macrophage differentiation was also studied. RESULTS: The co-culture of pMSCs with monocytes stimulated to follow the inflammatory M1 macrophage differentiation pathway resulted in a shift to anti-inflammatory M2-like macrophage differentiation. This transition was characterized by morphological of changes typical of M2 macrophages, and by changes in cell surface marker expression including CD14, CD36, CD163, CD204, CD206, B7-H4 and CD11b, which are distinctive of M2 macrophages. Co-culture with pMSCs reduced the expression of the costimulatory molecules (CD40, CD80 and CD86) and increased the expression of co-inhibitory molecules (CD273, CD274 and B7-H4) as well as the surface expression of major histocompatibility complex (MHC-II) molecules. Furthermore, the secretion of IL-10 was increased while the secretion of IL-1ß, IL-12 (p70) and MIP-1α was decreased; a profile typical of M2 macrophages. Finally, pMSCs induced the phagocytic activity and the phagocytosis of apoptotic cells associated with M2- like macrophages; again a profile typical of M2 macrophages. We found that the immunoregulatory effect of pMSCs on macrophage differentiation was mediated by soluble molecules acting partially via glucocorticoid and progesterone receptors. CONCLUSIONS: We have shown that pMSCs can transition macrophages from an inflammatory M1 into an anti-inflammatory M2 phenotype. Our findings suggest a new immunosuppressive property of pMSCs that may be employed in the resolution of inflammation associated with inflammatory diseases and in tissue repair.

Phenotypic and functional characterization of mesenchymal stem cells from chorionic villi of human term placenta.

BACKGROUND: Bone marrow derived mesenchymal stem cells (BM-MSCs) are used extensively in transplantation but their use is associated with many problems including low abundance in BM, low overall number, decreased differentiation potential with age and the invasive isolation procedures needed to obtain BM. We report a novel method of isolating placental MSCs (pMSCs) from chorionic villi, which exhibit the phenotypic and functional characteristics that will make them an attractive source of MSCs for cell-based therapy. METHODS: A novel explant approach was used to isolate pMSCs from chorionic villi of human placentae. These pMSCs were characterized by flow cytometry and were differentiated into adipocytes, osteocytes and chondrocytes using differentiation medium as demonstrated by cytochemical staining. The gene and protein expression profiles of pMSCs were also characterized using real time polymerase chain reaction (PCR) and flow cytometry, respectively. In addition, cytokine secretion by pMSCs was also analysed using sandwich enzyme-linked immunosorbent assay (ELISA) technique. Moreover, the migration and proliferation potentials of pMSCs were also determined. RESULTS: pMSCs were isolated from fetal part of the chorionic villi and these pMSCs expressed CD44, CD90, CD105, CD146, CD166 and HLA-ABC but not CD14, CD19, CD40, CD45, CD80, CD83, CD86 and HLA-DR. In addition, these pMSCs differentiated into osteocytes, chondrocytes and adipocytes and they also expressed several adhesion molecules, chemokines/receptors, growth factor receptors and cytokines/receptors. Moreover, they secreted many cytokines (IL-1Ra, IL6, IL8, IL10, IL11 and IL15) and they were able to proliferate. Furthermore, they migrated in response to chemotactic factors including stromal cell-derived factor-1 (SDF-1), platelet derived growth factor (PDGF), hepatocyte growth factor (HGF), and monocyte chemotactic protein-1 (MCP-1). CONCLUSIONS: We devised a novel explant method of isolating pMSCs that expressed many biological factors responsible for mediating cellular processes such as migration/homing, immune modulation and angiogenesis. Therefore, we suggest that pMSCs prepared from human term placental chorionic villous explants are an attractive source of MSCs for cell therapy.

Intrafamilial clustering of Helicobacter pylori infection in Saudi Arabia.

AIM: To study the pattern of Helicobacter pylori infection among family members in the Saudi population. METHODS: A cross-sectional, population-based, seroepidemiological study of family members was undertaken in a Saudi population using saliva H pylori immunoglobulin (Ig) G antibodies (Helisal kit). RESULTS: A total of 42 families comprising 271 children and 84 parents were studied (355 subjects; mean age 23 years, SD 19 years) The overall frequencies of H pylori IgG antibodies in mothers, fathers and children were 67%, 64% and 23%, respectively. There was no significant difference in the infection rate between mothers and fathers, or between boys and girls. The infection rate among children increased when one or both parents were seropositive, and the infection rate among parents was proportionally related to the number of infected children per family. The frequency of H pylori antibodies was significantly higher in spouses of seropositive parents than in spouses of seronegative parents (45% compared with 19.2%). CONCLUSIONS: These data confirm that the intrafamilial clustering of H pylori infection in Saudi Arabia occurs in a similar pattern to that described in the developed countries, and that living conditions and social conditions lead to person to person transmission of H pylori infection.

Hepatitis C Virus infection in Saudi Arab patients with B-cell non-Hodgkin's lymphoma.

OBJECTIVE: The objective of the current study is to determine the prevalence of Hepatitis C virus infection in Saudi Arab patients with B-cell non-Hodgkin's lymphoma. METHODS: Fifty-six unselected Saudi Arab patients with B-cell non-Hodgkin's lymphoma were tested for the presence of Hepatitis C virus antibodies using Elisa immunoabsorbant assay 2.0. Positive and indeterminate results were subjected to confirmatory testing using RIBA-Hepatitis C virus 2.0. Two control groups were utilized for comparison; the first is a group of randomly selected general medical patients and healthy blood donors; and the 2nd is a cohort of patients with hematological neoplasms other than B-cell non-Hodgkin's lymphoma. Patients with previous history of blood transfusion or liver disease were excluded from the study. RESULTS: Twelve of the 56 B-cell non-Hodgkin's lymphoma patients (21%) tested positive for Hepatitis C virus antibodies. Only 3 out of 104 (3%) and 2 out of 41 (5%) patients tested positive for Hepatitis C virus antibodies in the first and 2nd control groups. CONCLUSION: The results of this study indicate a higher prevalence of Hepatitis C virus infection in Saudi Arab patients with B-cell non-Hodgkin's lymphoma than in the control groups. The prevalence of Hepatitis C virus infection in the 2 control groups, in turn, seems to fall within the estimated prevalence in the general population.

Duodenal ulcer and Helicobacter pylori infection at high altitude: experience from southern Saudi Arabia.

OBJECTIVE: To study the clinical presentation, endoscopic features and prevalence of Helicobacter pylori in duodenal ulcer (DU) patients in southern Saudi Arabia, located 3150 m above sea level, and to compare results with those from low altitude regions of the Kingdom. METHODS: Prospective study of patients with proven DU referred for upper gastrointestinal endoscopy at Asir Central Hospital, Abha, southern Saudi Arabia over an 18-month period. RESULTS: Of 126 patients with proven DU, 72% were men and mean age was 40.4 years (range 18 to 68). Twenty-eight per cent were smokers and only 5% used nonsteroidal anti-inflammatory drugs. Thirty-eight patients (30%) presented with hematemesis or melena, and the majority had a single ulcer. Nineteen per cent of patients with dyspepsia had DU and 96% had H pylori. These results are comparable with those reported from the low altitude, warmer regions of Saudi Arabia. CONCLUSIONS: Age of patients and the male:female ratio were similar to those in developing countries. The frequency of smoking is lower than in western countries and no patient in this report consumed alcohol. High altitude did not affect the prevalence of DU or the frequency of H pylori because the results were comparable with those from the low altitude areas of the Kingdom of Saudi Arabia and other lowland developing countries. Although great socioeconomic changes have increased the incidence of heart disease, the patterns of DU and H pylori infection assume those in developing nations.

Non-neoplastic changes in gastric antrum: are they different in distally located intestinal and diffuse-type gastric adenocarcinoma?

A total of 126 cases of primary adenocarcinoma of distal (antrum and/or adjacent body) stomach were reviewed. These cases were collected from the histopathology laboratory of Asir Central Hospital, Southwestern Saudi Arabia over an 8 year period (1987-94). Only gastrectomy specimens with non-neoplastic antral mucosa available for histological examination were included. Of 126 cases, 85 (67.5%) were of the intestinal type and 41 (32.5%) were of the diffuse type. Histological examination of the non-neoplastic antral mucosa showed: gastritis in 100% of these cases; Helicobacter pylori in 103/126 cases (81.8%); multifocal atrophic gastritis (MAG) in 53/126 cases (42.1%); intestinal metaplasia (IM) in 62/126 (49.2%); and type III intestinal metaplasia in 30/62 cases (47.7%). None of these non-neoplastic changes of antral mucosa was significantly different when the prevalence of these changes in intestinal and diffuse type gastric adenocarcinoma were compared using the chi 2 test. The prevalence of these non-neoplastic lesions were calculated in a 126 dyspeptic age- and sex-matched control patients and were as follows: H. pylori 91%; gastritis 78%; MAG 7.4%; IM 19% and type III IM 1.6%. The prevalence of H. pylori bacilli and gastritis was not significantly different between the cancer patients and the controls. The prevalence of MAG, IM and type III IM was significantly higher among cancer patients compared with the control group.

Etiology of ascites and the diagnostic value of serum-ascites albumin gradient in non-alcohol liver disease.

This study was designed to determine the different etiologies of ascites and the diagnostic value of serumascites albumin gradient (SAAG) in patients with ascites of non-alcoholic liver disease in Southern Saudi Arabia. A total of 132 patients with ascites (96 males and 36 females, mean age 58.8+/-15.9 years) were studied for the different causes of ascites. In 55 patients with liver disease and 22 patients with nonliver disease (malignancy and peritoneal tuberculosis), we compared SAAG with the three usual parameters of ascitic fluid biochemical analysis used in the differential diagnoses of ascites. The nonliver disease group showed higher ascitic fluid total protein (aTP) concentration (4.77+/-2.05 versus 1.98+/-1.56 g/dL), ascitic to serum ratio of total protein (a/sTP) concentration (0.75+/-0.43 versus 0.26+/-0.19), ascitic fluid lactic dehydrogenase (aLDH) level (565.4+/-353.4 versus 254.1+/-205.03 U/L) and a lower SAAG (0.6+/-0.30 versus 1.71+/-0.61). P7lt;0.0001 for all parameters. The positive predictive values for aTP, a/sTP, aLDH and SAAG to detect ascites due to liver disease were 68%, 76%, 67%, and 80%, respectively, while the negative predictive values were 96%, 96%, 84%, and 98%, respectively. Liver causes accounted for 69.7% of cases, followed by peritoneal tuberculosis 10.6%, malignancy 9.1%, congestive heart failure 7.6%, and nephrotic syndrome 3.0%. SAAG is a useful diagnostic parameter which can be used to separate ascites of liver disease (nonalcoholic) from other causes of ascites, with an efficiency of 91%. SAAG should replace the traditional parameters (aTP, a/sTP, and aLDH) used in the differential diagnosis of ascites. In our series, liver disease is the major cause of ascites, followed by peritoneal tuberculosis.

A second-generation method of genotyping hepatitis C virus by the polymerase chain reaction with sense and antisense primers deduced from the core gene.

A second-generation method of genotyping hepatitis C virus (HCV) was developed by the polymerase chain reaction (PCR) with sense as well as antisense primers deduced from the core gene. HCV RNA specimens extracted from sera were reverse-transcribed and amplified with universal primers in the first round of PCR to obtain fragments of 433 base pairs representing nucleotides 319-751. In the second round of PCR, portions of PCR products were amplified separately with sense and antisense primers specific for each of the five common genotypes prevailing across the world, i.e., I/1a, II/1b, III/2a, IV/2b and V/3a. The specificity of the method was verified by a panel of 177 HCV isolates of various genotypes in the genetic groups 1-9. It allowed clear differentiation of genotype I/1a from II/1b which was not always accomplished by the previous method. When 501 sera from blood donors and hepatitis patients with HCV viremia from various countries were genotyped by the second-generation method, 478 (95.4%) were classified into the five genotypes. HCV RNA samples from 23 (4.6%) sera were not classifiable into any of the five common genotypes and, by sequence analysis, 22 were found to be of four genotypes in group 4 and one of genotype 1c in Simmond's classification.