Positive Cells In Patients Research Articles

Antimicrobial Peptides and Interleukins in Cleft Soft Palate.

Cleft palate is one of the most common and well-studied congenital anomalies; however, the role of protective tissue factors in its pathophysiology is still debated. The aim of our study was to evaluate interleukin and antimicrobial peptide appearance and distribution in cleft palate. Eight soft palate samples were obtained during veloplasty procedures. Immunohistochemical staining was applied to detect HBD-2-, HBD-3-, HBD-4-, LL-37-, IL-10-, and CD-163-positive cells via light microscopy. For statistical evaluation, the Mann-Whitney U test and Spearman's rank correlation coefficient were used. A significant difference between study groups was observed for HBD-2 and IL-10 in epithelial and connective tissue as well as HBD-4 in connective tissue. The number of HBD-3-positive cells was moderate in the patients, and few were observed in the controls. The number of LL-37-positive cells varied from a moderate amount to a numerous amount in both study groups, whilst CD-163 marked a moderate number of positive cells in patients, and a few-to-moderate amount was observed in the controls. Numerous correlations between studied factors were revealed in cleft tissues. The increase in antimicrobial peptides HBD-2 and HBD-4 and anti-inflammatory cytokine IL-10 suggested a wide compensatory elevation of the local immune system against cleft-raised tissue changes. The correlations between the studied factors (HBD-2, HBD-3, HBD-4, LL-37, and IL-10) proved the synergistic involvement of common local defense factors in postnatal cleft palate morphopathogenesis.

Evaluation of the Multiple Tissue Factors in the Cartilage of Primary and Secondary Rhinoplasty in Cleft Lip and Palate Patients.

Cleft lip and palate (CLP) is one of the craniofacial defects. The objective of this study was to identify the differences in appearance between the tissue factors in cartilage of CLP patients after primary and secondary rhinoplasty. Immunohistochemistry was performed with MMP-2, MMP-8, MMP-9, TIMP-2, IL-1α, IL-10, bFGF, and TGFβ1. The quantification of the structures was performed using a semi-quantitative census method. MMP-2, -9, IL-1a, and bFGF demonstrated higher number of positive cells in patients, while the number of MMP-8, IL-1a, -10 and TGFβ1 cells was higher or equal in the control subjects. The only statistically significant difference between CLP-operated patients was found in the TIMP-2 group, where the primary CLP patient group had a higher number of TIMP-2 positive chondrocytes than the secondary CLP patient group (U = 53.5; p = 0.021). The median value of the primary CLP group was ++ number of TIMP-2 positive chondrocytes compared to +++ in the secondary CLP group. No statistically significant difference was found between primary and secondary rhinoplasty patients for other tissue factors. Commonly, the rich expression of different tissue factors suggests a stimulation of higher elasticity in cleft affected cartilage. The statistically significant TIMP-2 elevation in primary operated cartilage indicates an impact of the selective tissue remodeling for hard tissue.

Heat shock protein gp96 and CD4+ and CD8+ T-lymphocytes expression as prognostic factors in various molecular types of invasive breast carcinoma.

Breast carcinoma shows extensive clinical and molecular heterogenicity. Glycoprotein gp96 is considered a negative prognostic and predictive factor. Controversy exists over the prognostic role of tumor lymphocytic infiltrates. The goal of this study is to illustrate differences in gp96 and CD4+ and CD8+ T-lymphocytes expression among all immunohistochemical groups of breast carcinoma in relation to the clinical course and outcome of the disease. A retrospective observational study was conducted through processing and analysis of 152 female patient tissue samples previously classified by immunohistochemistry. After immunohistochemical processing, the samples were microscopically analyzed and positive cells were manually calculated in the entire biopsy sample for each patient. In the group of patients with triple negative carcinoma, a significantly higher number of CD4 positive cells in patients with no local recurrence were proven, as well as a significant correlation between a smaller number of CD4 positive cells with a lethal outcome. In the group of patients with Luminal B HER2+ carcinoma, a significantly higher proportion of CD8+ cells in patients with local recurrence were demonstrated. The highest glycoprotein gp96 expression was demonstrated in the group of patients with triple negative carcinoma, while the lowest in patients with Luminal A and Luminal B HER2- carcinoma. This study has shown significantly higher gp96 expression and higher extent of tumor lymphocytic infiltrate in more malignant types of breast carcinoma and represents a significant contribution in affirmation of the prognostic role of these variables.

Abstract P3-04-01: Molecular determinants of post-mastectomy breast cancer recurrence

Abstract Introduction The management of breast cancer (BC) patients who undergo mastectomy in the setting of 1-3 positive lymph nodes has been controversial. This retrospective Translational Breast Cancer Research Consortium study evaluated the molecular aberrations associated with locoregional recurrence (LRR) or distant metastasis (DM) compared to controls in an effort to identify molecular predictors associated with recurrence. Methods/Materials We identified 115 HER2 negative, therapy naïve, T 1-3 and N 0-1 BC patients treated with mastectomy and no post mastectomy radiation therapy from 1997 to present with available FFPE tissue blocks. The cohort included 32 patients with LRR, 34 with DM, and 49 controls (without recurrence) who were matched for stage, grade, hormone receptor status, age ≤ or > 50, chemotherapy receipt, and margin status. Matched primary and recurrent LRR samples were available for 3 patients. Hybrid capture next generation sequencing (NGS) of 142 cancer related genes and RNAseq were performed to identify DNA/RNA alterations associated with LRR or DM. The frequency of common alterations on NGS was compared with Fisher's exact test. Expression of each gene from mRNA-Seq was treated as an explanatory variable. Immunohistochemistry (IHC) was performed for PTEN, Ki-67 and cleaved caspase 3 (CC3). PTEN loss and percentage of Ki-67 and CC3 positive cells were compared between groups with Fisher's exact test and nonparametric methods, respectively. Results RNAseq was performed on 115 patients; there was no difference in RNA expression levels between the groups. DNA analysis was performed on 57 patients (17 LRR, 15 DM and 25 controls), NF1 mutation rate was significantly elevated in both the LRR (24%) and DM (27%) samples compared to controls 0%; (p=0.0070). The mitogen activated protein kinase (MAPK) pathway was significantly mutated in both LRR (47%) and DM (40%) samples compared to the controls 0%; (p<0.0001). There was no significant difference in the rate of alterations of the PI3K/Akt/mTOR pathway among the three groups. Of three patients with matched primary vs LRR samples, one had concordant mutations. The second patient had additional mutations in the LRR, including gain of a NF1 mutation. The third patient had complete discordance of mutations identified in primary and LRR and had gain of HER2 amplification, suggestive of a new primary. There was no significant association between the groups and the loss of PTEN expression or CC3 expression. There was a significant difference between Ki 67 positive cells in patients with LRR (mean 29%), DM (mean 26%) versus controls (mean 14%, p= 0.0011). HR+ patients were significantly more likely to have a positive PTEN, lower Ki-67 and lower CC3 expression, p=0.0004, p<0.0001, and p<0.0001 respectively. Conclusions In this matched cohort analysis, mutations in the MAPK pathway, specifically NF1, were associated with both LRR and DM, suggesting that alterations in this pathway are associated with a more aggressive tumor phenotype. However, there were no molecular features that discriminated between those likely to recur locally alone versus distantly. Further study is needed to validate these findings, and to determine whether targeting alterations in this pathway could decrease the risk of recurrence. Citation Format: Keene KS, King T, Hwang ES, Peng B, McGuire K, Tapia C, Zhang H, Bae S, Nakhlis F, Klauber-Demore N, Meszoely I, Sabel MS, Willey SC, Eterovic KA, Hudis C, Wolff A, De Los Santos J, Thompson A, Mills GB, Meric-Bernstam F. Molecular determinants of post-mastectomy breast cancer recurrence [abstract]. In: Proceedings of the 2017 San Antonio Breast Cancer Symposium; 2017 Dec 5-9; San Antonio, TX. Philadelphia (PA): AACR; Cancer Res 2018;78(4 Suppl):Abstract nr P3-04-01.

The role of adiponectin gene mediated by NF-κB signaling pathway in the pathogenesis of type 2 diabetes.

To investigate the role of adiponectin (ADPN) gene mediated by NF-κB signaling pathway in the pathogenesis of type 2 diabetes (T2DM). The levels of expression of ADPN were measured by fluorescence quantitative PCR technology, enzyme-linked immunosorbent assay, Western-blotting and immunohistochemistry in 78 patients with type 2 diabetes. Compared with the normal population, the levels of expression of ADPN and NF-κB mRNA in the blood of patients with type 2 diabetes were significantly reduced (p < 0.05). The detection of ADPN gene protein expression in normal population and patients with type 2 diabetes by enzyme-linked immunosorbent assay showed that ADPN gene protein expression in type 2 diabetic patients (1.26 ± 0.73) µg/l was significantly lower (p < 0.05) than ADPN gene protein expression of the normal population (3.26 ± 1.25) µg/l. At the same time, the detection of the expression of NF-κB gene showed that the protein expression in type 2 diabetic patients (0.58 ± 0.15) µg/l was significantly lower (p < 0.05) than that in the normal population (1.67 ± 1.04) µg/l. The results of Western-blotting were consistent with the results of enzyme-linked immunosorbent assay. Immunohistochemical results also showed that the percentage of ADPN positive cells in patients with type 2 diabetes (25.47%) was significantly lower (p < 0.05) than that in the normal population (79.47%). ADPN gene in the human body can be involved in the pathogenesis of T2DM through the NF-κB signaling pathway.

Preclinical characterisation of the GM-CSF receptor as a therapeutic target in rheumatoid arthritis

ObjectivePrevious work has suggested that the granulocyte macrophage colony stimulating factor (GM-CSF)–GM-CSF receptor α axis (GM-CSFRα) may provide a new therapeutic target for the treatment of rheumatoid arthritis (RA). Therefore,...

Intrathecal Administration of Autologous CD34 Positive Cells in Patients with Past Cerebral Infarction: A Safety Study

Regenerative strategies in treatment of stroke have great potential. The goal of the current study was to investigate safety of intrathecal administration of autologous CD34 positive cells in treatment of patients with poststroke. A total of eight male patients with a history of stroke were enrolled. The patients were treated subcutaneously with 5 μg/kg body weight rhG-CSF for 5 consecutive days, and then leukapheresis was performed to concentrate cells for CD34 positive immunoselection. All patients underwent intrathecal administration of CD34 positive cells via lumbar puncture. The primary outcome was safety evaluation for 12-month followup. In addition, behavioral function was evaluated with NIH stroke scale and Barthel index 1, 6, and 12 months after the last treatment, respectively. There were no major adverse events, and abnormal changes of blood tests during the whole treatment process included intrathecal administration and 12-month followup. The main message from the current study was that administration of G-CSF-mobilized autologous CD34 positive cells in patients with poststroke was safe. Future studies with larger population and control group are needed to confirm the safety and investigate the efficacy.

Abstract P5-03-13: Detection of tumor initiating cells (TIC) among the peripherally circulating epithelial tumor cells from patients with breast cancer

Abstract Background: The detection of tumor cells circulating in the peripheral blood of patients with breast cancer is a sign that cells have been able to leave the primary tumor and survive in the circulation. However, in order to form metastases they require additional properties such as the ability to adhere, self renew and grow. Here we present data that a variable fraction among the circulating tumor cells detected by the maintrac® approach express mRNA of the stem cell gene nanog and of the adhesion molecule vimentin and are capable of forming tumor spheres a property ascribed to TIC. Patients and methods: Between 10 to 50 circulating epithelial antigen positive cells detected by the maintrac® approach were selected randomly from each of 20 patients with breast cancer before and after surgery and isolated using automated capillary aspiration and deposited individually onto slides for expression profiling. In addition, the circulating tumor cells were cultured without isolation among the white blood cells from 5 patients with breast cancer in different stages of disease using culture methods favoring growth of epithelial cells. Results: Although no EpCAM mRNA positive cells expressing stem cell genes or the adhesion molecule vimentin were detected before surgery, 10 to 20% of the cells were found to be positive for mRNA of these genes after surgery. Most surprisingly, it was possible to grow tumor spheres (Fig 1) from these circulating cells without previous isolation in all patients in a fraction comprising between 1% of the epithelial antigen positive cells in patients with newly diagnosed tumor up to 10% in a patient with advanced breast cancer. Conclusion: We here show, that among the peripherally circulating tumor cells a variable fraction is able to express stem cell and adhesion properties and can be grown into tumor spheres, a property ascribed to cells capable of initiating tumors and metastases. Citation Information: Cancer Res 2012;72(24 Suppl):Abstract nr P5-03-13.

BCL2L10 (Bcl-B) Is Associated with Resistance to Azacitidine (AZA) in MDS and AML, and Is a Possible Therapeutic Target in AZA Resistant Patients

AbstractAbstract 701 Background:AZA is a current reference treatment for higher-risk MDS patients, also active in AML patients unfit for intensive chemotherapy. However, no biological marker predictive of AZA resistance has been clearly established. In addition, patients not responding to AZA or relapsing after a response have very poor outcome (Prebet, JCO, 2011), and require new treatments. We have generated SKM1-R, an AZA resistant cell line (Cluzeau et al., Cell cycle 2011), that exhibits increased expression of BCL2L10, an anti-apoptotic Bcl-2 family member, as a likely cause of resistance (Yasui et al., Cancer Research 2004). We tried to correlate BCL2L10 expression with response to AZA in MDS and AML patients and analysed the effect of ABT 737 a peptidomimetic inhibitor binding the BH3 domain of the BCL-2 family of proteins (especially BCL2L10), on patient leukemic cells expressing BCL2L10. Methods:In two cohorts of 77 MDS or AML patients treated by AZA (75mg/m2/d, 7days every 4 weeks), we quantified the percentage of BCL2L10 positive bone marrow (BM) cells using flow cytometry. Cytometry assay was performed after several steps of fixation, permeabilization, primary incubation with an anti-BCL2L10 antibody (cell signalling) and secondary incubation with a donkey anti-Rabbit FITC-antibody. The first cohort included fresh BM samples from 32 higher-risk MDS (www.clinicaltrials.gov, NCT01210274) or AML patients treated with AZA. The second one is composed of 45 frozen samples from low risk MDS, higher-risk MDS or AML treated by AZA. Response and relapse after AZA initial response were assessed by IWG 2006 criteria. Cell metabolism (XTT assay) and Propidium iodide (PI) staining were performed in 10 patients treated with AZA (5 sensitive and 5 resistant) with or without ABT-737 (Abbott). Resistance to AZA was defined by primary failure or relapse after initial sensitivity to AZA Results:In the first patient cohort, the mean value for BM cells from AZA-resistant patients was significant higher than AZA-sensitive patients (85% vs 0% respectively, p<0.0001). In the second cohort, retrospective comparison of BM samples from low risk MDS patients, AZA-sensitive or AZA-resistant patients showed that the counts of BCL2L10 positive cells were also significant different (0%, 10% and 33% respectively, p<0.0001). In sub-group analysis on patients from cohort 2, we observed that quantification of BCL2L10 positive cells could predict failure to AZA at diagnosis (10% in sensitive patients vs 33% in resistant patients, p=0.023) and relapse after response to AZA (14% in AZA responding patients without relapse vs 43% in AZA responding patients who relapsed, p=0.0002). Using a cut-off of 50% BCL2L10 positive cells in patients from cohort 1, patients with > 50% of BCL2L10 positive cells had a 3 months OS at 51% compared to 95% in patients with ≤ 50% of BCL2L10 positive cells (p=0.0016).We next used high doses of ABT-737, alone or in combination with AZA. Cell metabolism (XTT assay) and Propidium iodide (PI) staining in the presence or absence of ABT 737 showed no effect in AZA-sensitive patients. By contrast, we observed a significant increase of PI positive cells and a significant decrease of cell metabolism in AZA-resistant patient samples stimulated with the combination of AZA + ABT-737 vs AZA alone. Conclusion:In this work, quantification of BCL2L10 positive cells by flow cytometry was correlated with resistance and survival with AZA treatment in MDS and AML. This possible biomarker of resistance will have to be tested prospectively. Inhibition of BCL2L10 positive cells by ABT-737 suggests that this drug, or other drugs targeting more selectively BCL2L10 could be proposed in combination with AZA in such patients. Disclosures:No relevant conflicts of interest to declare.

Distinct expression of interleukin 17, tumor necrosis factor α, transforming growth factor β, and forkhead box P3 in acute rejection after kidney transplantation

The kidney transplant is the main therapeutic alternative for end-stage kidney disease, and rejection is a major complication. The expression of proinflammatory cytokines is related to graft loss, whereas anti-inflammatory cytokines are associated with graft protection. The objective of this study is to evaluate the "in situ" expression of cytokines T helper 1 (tumor necrosis factor α [TNF-α]), T helper 17 (interleukin 17 [IL-17]), and regulatory T cell (transforming growth factor β [TGF-β]) and the expression of forkhead box P3 (FoxP3) in allograft kidney. We evaluated in situ expression of cytokines in allograft kidney under rejection process by indirect immunohistochemistry. Eighteen renal graft biopsies were from patients with episodes of rejection. The in situ expression of IL-17, TNF-α, and TGF-β was significantly higher in patients with acute rejection when compared with the control group. In contrast, analysis of FoxP3 expression showed few positive cells in patients with acute rejection compared with the control group. The results suggest that the expression of proinflammatory cytokines (IL-17 and TNF-α) contributes to the mechanisms of kidney transplant rejection. The increase in TGF-β expression might be an attempt to establish a process of immunoregulation or even to induce higher production of IL-17. The last hypothesis is supported by the observation of a reduced expression of FoxP3 and elevated levels of IL-17.

Long-term clinical outcome after intramuscular transplantation of granulocyte colony stimulating factor-mobilized CD34 positive cells in patients with critical limb ischemia

BackgroundOur phase I/IIa clinical trial revealed that intramuscular transplantation of autologous, GCSF-mobilized CD34+ cells was safe, feasible and potentially effective at week 4 and 12 post cellular therapy in 17 patients with chronic critical limb ischemia (CLI) (5 patients with atherosclerotic peripheral arterial disease (PAD) and 12 with Buerger's disease). However, long-term outcome of the cell therapy has yet to be reported. Methods and resultsIncidence of major clinical events and physiological parameters of limb ischemia were evaluated at week 52, 104, 156 and 208 post CD34+ cell therapy. No patients died by week 104, whereas 3 patients with PAD died by week 156 and 1 patient with Buerger's disease died by week 208 due to cardiac complications. No patients underwent major amputation, whereas 1 patient with Buerger's disease underwent unplanned minor amputation by week 104. CLI-free ratio was 88.2% at week 52 and 104, 92.3% at week 156 and 84.6% at week 208 in all patients. Significant improvement of toe brachial pressure index versus baseline was sustained up to week 208 and that of transcutaneous partial oxygen pressure was kept up to week 156. The Wong-Baker FACES pain rating scale, ulcer size and exercise tolerance significantly improved at week 52, the final evaluation time point, compared with baseline. Subgroup analysis revealed the similar outcome in patients with Buerger's disease. ConclusionsFavorable clinical outcomes as well as physiological evidences strongly indicate the long-term benefit of GCSF-mobilized CD34+ cell transplantation for retrieval from CLI, especially in patients with Buerger's disease.

GATA Transcription Factors and Hematological Diseases

The development of mature blood cells from hematopoietic stem cells is regulated by transcription factors that control and coordinate the expression of lineage-specific genes. The GATA family consists of six transcription factors that function in hematopoietic and endodermal development. Among them, GATA-1 is expressed in erythroid, megakaryocytic, eosinophil and mast cell lineages, and GATA-2 is expressed in stem and progenitor cells, at more immature stage compared with GATA-1. Based on the characteristic phenotypes of GATA-1 and GATA-2 mutant mice, it has been suggested that mutations of these GATA genes in humans may result in the onset of certain clinical diseases. To date, mutations of GATA-1 gene have been found in inherited anemia and thrombocytopenia, and Down syndrome-related acute leukemia, which exhibits megakaryocytic phenotypes and frequently occurs in patients with Down syndrome. In contrast, no mutation of GATA-2 gene has been identified in hematological diseases; however, we found the expression level of GATA-2 is significantly decreased in CD34 positive cells in patients with aplastic anemia. Since GATA-2 functions in the proliferation of hematopoietic stem cells, the reduction of GATA-2 expression in CD34 positive cells may result in the decreased number of hematopoietic stem cells, which is the characteristic feature of aplastic anemia. Based on these lines of evidence, some types of hematological diseases may be defined as transcription factor diseases.

Molecular detection of HER2 mRNA-positive cells in the peripheral blood of patients with operable breast cancer

668 Background: To evaluate whether the detection of peripheral blood (PB) HER2 mRNA+ positive cells in patients with breast cancer could be another marker of the micrometastatic disease. Methods: The sensitivity and specificity of HER2 mRNA detection by nested RT-PCR were investigated using MCF-7 and ARH-77 cells as well as blood from healthy women and patients with colorectal, early and metastatic breast cancer. PB from 216 patients with operable breast cancer, obtained before initiation of any adjuvant therapy, was tested for the presence of HER2 mRNA+ positive cells. Results: In dilution experiments the nested RT-PCR assay for HER2 mRNA was capable of detecting up to 105 MCF-7 tumor cells in 105 normal peripheral blood mononuclear cells; no signal was detected with the HER2- ARH-77 cells. No HER2 mRNA+ cells could be detected in the peripheral blood of 31 healthy women, as well as the peripheral blood and the bone marrow of 20 patients with colorectal cancer. Detection rates for HER2 mRNA+ cells in the bone marrow/blood of patients with early or metastatic breast cancer were 36.4%/23.8% and 35.5%/18%, respectively. Double immunostaining with a pan-keratin and a HER2 monoclonal antibody and confocal microscopy analysis revealed co-expression on PB cells in 5 (38.5%) of 13 patients. In addition, FISH analysis of PB tumor cells from 5 patients demonstrated HER2 gene amplification in all patients. Finally, aneusomy cells could be detected by LSI 1, CEP 8, CEP 11 and CEP 11 with chromosomal gain or loss in 2 of the patients. Conclusions: The results demonstrate that occult tumor cells expressing HER2 mRNA could be detected in the peripheral blood of patients with operable breast cancer; these cells are not always CK+ indicating that HER2 mRNA detection may represent another marker of the micrometastatic disease. No significant financial relationships to disclose.

Peri-operative filtration of disseminated cytokeratin positive cells in patients with colorectal liver metastasis

Liver resection is the only potential cure for patients with colorectal liver metastasis. However, more than 30% of patients will develop tumour recurrence, probably caused by tumour cells disseminated before or during surgery. As prevention of cell dissemination is barely obtainable, alternative concepts have to be discussed. The potential of leukocyte adhesion filters for the removal of cytokeratin positive cells (CK+) from blood was studied in 18 patients undergoing liver resection for colorectal liver metastasis. Blood sampling was done via a liver venous catheter during hepatic mobilisation. Filtration was done with an in-line WBF2 filter system. To define the relation between surgery and cell release we compared patients' pre-operative and intra-operative blood and bone marrow (BM) samples with their CK expression using immunochemical staining. CK+ cells were detected in BM samples of nine of 14 patients before surgery, indicating early dissemination. In ten of 18 patients CK+ cells were detected in blood samples during hepatic mobilisation; all ten patients underwent major liver surgery (R0 resection). In those patients recurrent disease was observed more often (P < or = 0.05). In 17 of 18 patients CK+ cells were not detectable after filtration procedure, which indicated cell adhesion to the filter medium. Liver resection due to metastasis leads to frequent intra-operative tumour cell shedding. As the detection of CK+ cells is correlated with disease recurrence, modification of surgical techniques to prevent cell dissemination, and additional therapeutic concepts such as advanced filtration technology, have to be discussed.

Hepatic stellate cells in hepatitis C patients: relationship with liver iron deposits and severity of liver disease.

To determine the relationship between hepatic stellate cell (HSC) populations and severity of liver disease and liver iron deposits in patients with chronic hepatitis C virus (HCV). We also studied the relationship between iron cellular distribution and HSC population and the role of HFE mutations in the determination of iron deposits. Forty-nine chronic HCV patients with varying degrees of liver damage and liver iron deposits were studied. A liver biopsy was scored for histology activity index (HAI), fibrosis and iron deposits. The number of HSC in the liver was evaluated by an immunohistochemical double-staining method to identify glial fibrillary acid protein (GFAP) and smooth muscle alpha-actin (alpha-SMA). The HSC population was significantly higher in HCV patients than in normal controls and was predominant in zones 1 and 3. Liver iron deposits were observed in 49% of patients and were mild/moderate in most cases. We found a significantly higher number of GFAP and alpha-SMA positive cells in patients with liver iron deposits compared with those without iron deposits, and a positive correlation between liver iron scores and number (%) of GFAP and alpha-SMA positive cells. We observed a significantly higher number of GFAP and alpha-SMA positive cells in moderate/severe hepatitis than in minimal/mild hepatitis, and a positive correlation between GFAP and alpha-SMA positive cells and HAI and fibrosis scores. Liver iron deposits in chronic HCV are common and are associated with activation of HSC. Thus, even mild iron deposits might stimulate HSC and contribute to liver damage.

Intracellular cytokine profile of CD14 positive cells in patients with hematologic malignancies and solid tumors during hematologic recovery phase after intensive chemotherapy designed to mobilize peripheral blood stem cells

We studied intracellular cytokines in monocytes by flow cytometry from 28 patients with hematologic malignancies and solid tumors to analyze the role of monokines in the hematologic recovery phase for peripheral blood stem cell harvest. The patients were divided into three groups: the first group, A, had a documented infection; the second group, B, had fever of unknown origin; and the third group, C, was afebrile. We found an increase in intracellular IL-1α, IL-6, IL-8 and TNF-α positive monocytes as CD14 positive gated cells cultured with lipopolysaccharide in all groups, but no increase was found with medium only when cultured for 4 h. We also found an increase in intracellular IL-1a, IL-6, IL-8 and TNF-α positive monocytes cultured with autologous serum for 4 h, but only in group A. The rate of intracellular cytokine positive cells was higher in monocytes cultured with only autologous serum from group A patients compared to those cells from the other groups; the data concerning IL-1a, IL-6 and TNF-α reached statistical significance ( P < 0.05). However, increasing intracellular cytokine levels in the control group of patients exhibiting only infectious disease were observed. Thus, it appear that pro-inflammatory intracellular cytokine levels in monocytes are only related to microbial infections.

No evidence for persistence of BCR-ABL–positive cells in patients in molecular remission after conventional allogeneic transplantation for chronic myeloid leukemia

Appropriate monitoring of patients after allogeneic stem cell transplantation (SCT) for chronic myeloid leukemia (CML) is crucial, because the response to donor lymphocyte infusions is superior in the case of cytogenetic or molecular relapse compared with hematological relapse.[1][1] Chomel et al

Increased numbers of dendritic cells in the bronchiolar tissues of diffuse panbronchiolitis.

Dendritic cells (DCs) are potent antigen-presenting cells (APCs); they are considered to be the most important APC in the lung. Recently, the number of DCs in the large airways was demonstrated to increase in patients with atopic asthma, leading to the concept that DCs play an important role in airway inflammation. However, little is known about the distribution of lung DCs in the small airways under other pathological conditions. The aim of the present study was to examine the distribution of DCs in the bronchiolar tissues in patients with diffuse panbronchiolitis (DPB), which is a chronic inflammatory disorder of the airways histologically characterized by peribronchiolitis. We investigated the distribution of DCs in the bronchiolar tissues of the lungs in 11 patients with DPB and 7 control subjects with normal lungs using immunohistochemical methods. Marked increases in the number of CD1a(+), CD1c(+), and CD83(+) DCs were found in both the bronchiolar epithelium and submucosal tissues of patients with DPB, compared with control subjects with normal lungs. The most striking increase occurred in the number of DCs expressing CD83, a marker of mature DCs, in the submucosal tissues of patients with DPB. The increases of these positive cells in patients with DPB were more marked in the submucosal tissues than in the epithelium. The bronchiolar epithelial cells in patients with DPB strongly expressed GM-CSF protein, which is an important cytokine for the differentiation and function of DCs, suggesting that the increased local production of GM-CSF may be responsible for the accumulation and differentiation of DCs in the bronchiolar tissues of patients with DPB. These results suggest that increased DCs in the bronchiolar tissues, together with their phenotypical maturation, may play an important role in the mucosal immune response in patients with DPB through their potent antigen-presenting function.

Quantitative detection of t(14;18)-positive cells in patients with follicular lymphoma before and after autologous bone marrow transplantation.

The aim of this study was to evaluate whether a quantitative analysis of circulating t(14;18)-positive cells is of prognostic significance in patients with follicular lymphoma (FL) after myelo-ablative therapy supported by ABMT. We tested DNA from primary lymphoma tissue as well as PBMC before and after ABMT from 15 patients for the presence of the t(14;18) translocation. Nine patients showed a t(14;18) translocation, six patients were t(14;18)-negative. Circulating t(14;18)-positive cells of seven patients were quantitatively determined by limiting dilution assays combined with a two-step PCR and by real-time quantitative PCR. The results of both methods correlate very well. The number of circulating t(14;18)-positive cells decreased significantly in all patients after myeloablative therapy and ABMT, t(14;18)-negative blood samples were found in five of seven patients. In all patients circulating t(14;18)-positive cells reappeared within 2 years after ABMT showing two different patterns. During continuous CR the numbers of circulating t(14;18)-positive cells were found to be stable within one order of magnitude. In contrast, in one patient the relapse was accompanied by a logarithmic increase of t(14;18)-positive cells. In a second patient the enlargement of lymph nodes developing over a period of 12 months was accompanied by very slowly increasing numbers of t(14;18)-positive cells. In all cases where diagnostic lymph node tissue was available, the same t(14;18) translocation was found at first diagnosis and after ABMT as shown by nucleotide sequence analysis. We conclude that the quantitative detection of circulating t(14;18)-positive cells during follow-up of patients with FL after ABMT reflects the clinical course of the disease. Relapses are associated with increasing numbers of circulating t(14;18)-positive cells and continuous complete remissions with stable cell counts.

Dysregulation of Membrane-Bound Tumor Necrosis Factor-α and Tumor Necrosis Factor Receptors on Mononuclear Cells in Human Immunodeficiency Virus Type 1 Infection: Low Percentage of p75-Tumor Necrosis Factor Receptor Positive Cells in Patients With Advanced Disease and High Viral Load

The correlation of persistent tumor necrosis factor-α (TNF-α) activation with disease progression in patients infected with human immunodeficiency virus type 1 (HIV-1), suggests a role for TNF-α in the pathogenesis of HIV-1 infection. In the present study, we examined by flow cytometry the expression of membrane-bound (m) components of the TNF system in 33 HIV-1–infected patients and 12 healthy controls. While peripheral blood mononuclear cells (PBMC) from asymptomatic and symptomatic non-acquired immune deficiency syndrome (AIDS) patients showed a significantly increased percentage of mTNF-α+ and mTNF receptor (TNFR)+ cells compared with controls, this was not found in the AIDS group. Compared with healthy controls, AIDS patients had a significantly decreased percentage of both monocytes and lymphocytes expressing p75-TNFR. PBMC from AIDS patients showed a higher p75-TNFR mRNA level and a higher spontaneous release of soluble p75-TNFR than healthy individuals, suggesting enhanced cell surface turnover of this TNFR. The low expression of TNFRs on both lymphocytes and monocytes in the AIDS group was associated with high numbers of HIV-1 RNA copies in plasma, low numbers of CD4+ lymphocytes, and high serum levels of soluble TNFRs. AIDS patients had a decreased percentage of CD8+ lymphocytes expressing TNFRs compared with healthy controls. In contrast, these patients, as well as symptomatic non-AIDS patients, had an increased percentage of TNF-α+ and TNFRs+ cells among remaining CD4+ lymphocytes. The pattern of abnormalities seen in AIDS patients suggests a role for persistent activation of the TNF system in the accelerated CD4+ lymphocyte destruction, the enhanced HIV-1 replication, and the markedly impaired antimicrobial defense in advanced HIV-1-related disease.