Circulating Immune Cells Predict Prognosis and Clinical Response to Chemotherapy in Cholangiocarcinoma.

BACKGROUND: The immune system is linked to the prognosis and response to treatment of patients with cancer. However, the clinical implication of peripheral blood immune cells in cholangiocarcinoma (CCA) remains vague. Thus, we aimed to assess whether peripheral circulating immune cells could be used as an indicator for prognosis and chemotherapeutic efficacy in CCA. METHODS: The distributions of immune subsets were analyzed in peripheral blood samples from 141 patients with CCA and 131 healthy volunteers by using flow cytometry. The variation in the subset distribution in the two groups and the relationship between clinicopathological features and the subpopulations were investigated. Meanwhile, we assessed the implications of lymphocyte subsets as predictors of chemotherapy outcomes and overall survival (OS). RESULTS: The proportion of total lymphocytes decreased, while the percentages of activated T cells as well as CD4+CD25+ regulatory T cells (Tregs) increased in CCA. Notably, lymphocyte proportion decreased in patients with regional lymph node (N) (p=0.016) and distant metastasis (M) (p= 0.001). Furthermore, our study showed that peripheral blood lymphocyte subsets were significantly correlated with chemotherapy efficacy, with increased proportions of CD3+ cells (p=0.021) and CD4+ cells (p=0.016) in the effective group. Finally, the Kaplan-Meier analysis indicated that patients with high natural killer (NK) cell proportion might have prolonged OS (p = 0.028). CONCLUSION: The relationship between circulating immune cells with prognosis and chemotherapy response in patients with CCA highlights their potential application as an indicator of CCA prognosis and stratification of chemotherapy response.

Novel PBMC LncRNA signatures as diagnostic biomarkers for colorectal cancer.

The expression of long non-coding RNAs (LncRNAs) in peripheral blood mononuclear cell (PBMC) and its clinical relevance in colorectal cancer (CRC) remains largely uncharacterized. To address these gaps, we investigated the expression profiles of lncRNAs in PBMC from CRC and healthy controls (HC) by RNA sequencing. The expression level of differentially expressed lncRNAs (DElncRNAs) were evaluated by quantitative PCR in PBMC samples from CRC patients and HC. A total of 447 DElncRNAs were identified, with 178 elevated lncRNAs and 269 decreased lncRNAs in PBMC from CRC patients as compared with that from HC. RT-PCR results supported a significant elevation of NEAT1:11, lnc-PDZD8-1:5 and LINC00910:16 in 98 CRC patients and 82 HC. The clinical implication of NEAT1:11, lnc-PDZD8-1:5 and LINC00910:16 as CRC diagnostic biomarker were determined by receiver operating characteristic (ROC) curve, showing sensitivity 74.5% and specificity 84.5% for joint detection the three lncRNAs. Notably, NEAT1:11 was closely related with the size and extent of primary tumor, with higher relative expression of NEAT1:11 in higher T stage (P = 0.0047). Moreover, NEAT1:11 was related with grade (P = 0.012). Collectively, PBMC from patients with CRC show significantly variable expression profiles of lncRNAs, and detection of these differential expression lncRNAs may provide useful information for basic and clinical research.

Nanoproteomics deciphers the prognostic value of EGFR family proteins-based liquid biopsy.

Monitoring tumor-associated protein status in serum can effectively track tumors and avoid time-consuming, costly, and invasive tissue biopsy. Epidermal growth factor receptor (EGFR) family proteins are often recommended in the clinical management of multiple solid tumors. However, the low-abundance of serum EGFR (sEGFR) family proteins hinders the depth-understanding of their function and tumor management. Herein, a nanoproteomics approach coupling with aptamer-modified MOFs (NMOFs-Apt) with mass spectrometry was developed for the enrichment and quantitative analysis of sEGFR family proteins. This nanoproteomics approach exhibited high sensitivity and specificity for sEGFR family protein quantification, with the limit of quantification as low as 1.00 nM. After detecting 626 patients' sEGFR family proteins with various malignant tumors, we concluded that the levels of serum proteins had a moderate concordance with tissue counterparts. Metastatic breast cancer patients with a high level of serum human epidermal growth factor receptor 2 (sHER2) and a low level of sEGFR had a poor prognosis, and patients with a sHER2 decrease of more than 20% had longer disease-free time after receiving chemotherapy. This nanoproteomics method provided a simple and effective approach for low-abundant serum protein detection and our results clarified the potential of sHER2 and sEGFR as cancer markers.

Single-cell metabolic fingerprints discover a cluster of circulating tumor cells with distinct metastatic potential.

Circulating tumor cells (CTCs) are recognized as direct seeds of metastasis. However, CTC count may not be the "best" indicator of metastatic risk because their heterogeneity is generally neglected. In this study, we develop a molecular typing system to predict colorectal cancer metastasis potential based on the metabolic fingerprints of single CTCs. After identification of the metabolites potentially related to metastasis using mass spectrometry-based untargeted metabolomics, setup of a home-built single-cell quantitative mass spectrometric platform for target metabolite analysis in individual CTCs and use of a machine learning method composed of non-negative matrix factorization and logistic regression, CTCs are divided into two subgroups, C1 and C2, based on a 4-metabolite fingerprint. Both in vitro and in vivo experiments demonstrate that CTC count in C2 subgroup is closely associated with metastasis incidence. This is an interesting report on the presence of a specific population of CTCs with distinct metastatic potential at the single-cell metabolite level.

CircMERTK modulates the suppressive capacity of tumor-associated macrophage via targeting IL-10 in colorectal cancer.

Macrophages represent the major population in the tumor microenvironment (TME). Recent studies have demonstrated circular RNAs (circRNAs) are implicated in the development and progression of different immune responses and immune diseases. However, the role of circRNAs in the development of tumor-associated macrophages (TAM) remains unknown. Here, we used the circRNA sequencing to identify the differentially expressed circRNAs (DEcircRNAs) in TAM-like cell induced by culture medium of colorectal cancer cell lines. Of note, the expression of circMERTK was remarkably overexpressed in TAMs. The ISH assay displayed that the expressions of circMERTK were mainly overlapped with macrophages marker CD68, and the abundance of circMERTK in CRC tissues was much higher than that in matched normal tissues. Functionally, circMERTK knockdown resulted in attenuated CD8+ T cell apoptosis in the co-culture assay, indicating that circMERTK could have an impact on the immunosuppressive activity of TAM-like cell. Mechanically, TAM-like cell could exert immunosuppressive activity via circMERTK/miR-125a-3p/IL-10 axis. These data suggested that circMERTK could play an important role in TAM activation, and may serve as a potential therapeutic target for CRC.

Clinicopathological and prognostic significance of circulating immune cells in the patients with pancreatic cancer.

Pancreatic cancer is characterized by immune tolerance and immunotherapeutic resistance. Circulating cells may reflect the general immune status of the patient. However, the circulating immune status of pancreatic cancer are largely uncharacterized. Here, the subset distribution was analyzed in peripheral blood samples from 101 patients with pancreatic cancer and 142 healthy volunteers by using flow cytometry. The differences of the subpopulation distribution in the two groups and the relation between clinical parameters with the subset were determined. Moreover, the clinical application of each subset as prognosis biomarker was also assessed by Kaplan-Meier analysis. The reduced proportion of total lymphocyte and upregulated CD4/CD8 ratio were observed in pancreatic cancer than those in healthy controls. Of note, increased proportions of lymphocyte and NKT cells were noticed more frequently in patients over 60 years (P = 0.043) and patients with metastasis (P = 0.027), respectively. However, our correlation analyses revealed no correlation between the proportions of T cells, B cell and NK cells with clinicopathologic features. Furthermore, the analysis displayed that proportions of CD4+T cell, B cell and CD4/CD8 ratio significantly reduced in the cohort of post-operation, while the frequency of CD8+T cell and NKT cells elevated remarkably. Finally, the Kaplan-Meier analysis indicated that patients with high lymphocyte proportion might have prolonged overall survival (P = 0.007). The altered distribution of peripheral blood immune cell subpopulation in pancreatic cancer and its relationship with clinical outcome highlight the potential use of circulating immune subsets as prognostic biomarkers in pancreatic cancer.

The yin and yang functions of extracellular ATP and adenosine in tumor immunity.

Extracellular adenosine triphosphate (eATP) and its main metabolite adenosine (ADO) constitute an intrinsic part of immunological network in tumor immunity. The concentrations of eATP and ADO in tumor microenvironment (TME) are controlled by ectonucleotidases, such as CD39 and CD73, the major ecto-enzymes expressed on immune cells, endothelial cells and cancer cells. Once accumulated in TME, eATP boosts antitumor immune responses, while ADO attenuates immunity against tumors. eATP and ADO, like yin and yang, represent two opposite aspects from immune-activating to immune-suppressive signals. Here we reviewed the functions of eATP and ADO in tumor immunity and attempt to block eATP hydrolysis, ADO formation and their contradictory effects in tumor models, allowing the induction of effective anti-tumor immune responses in TME. These attempts documented that therapeutic approaches targeting eATP/ADO metabolism and function may be effective methods in cancer therapy.

Long Noncoding RNA TUG1 Promotes Autophagy-Associated Paclitaxel Resistance by Sponging miR-29b-3p in Ovarian Cancer Cells.

PURPOSE: Paclitaxel (PTX) is a first-line chemotherapeutic agent for treating ovarian cancer. However, PTX resistance has become a major obstacle in ovarian cancer therapy. The underlying mechanism associated with PTX resistance is still unclear. PATIENTS AND METHODS: We used qPCR to detect taurine up-regulated 1 (TUG1) expression in normal ovarian tissues and ovarian tumor tissues. A combination of small interfering RNA (siRNA), cell counting kit 8 (CCK8), colony formation assay and nude mouse model were used to detect the effect of TUG1 on ovarian cancer cell PTX-resistance. Autophagy/cytotoxicity dual staining assay, luciferase reporter assay, Western blot and RNA-binding protein immunoprecipitation assay were used for further mechanistic studies. RESULTS: TUG1 is highly expressed not only in ovarian tumor tissues compared with normal ovarian tissues but also in the chemo-resistant group compared with the sensitive group. Knockdown of TUG1 by siRNA decreased ovarian cancer cell and xenograft tumor PTX resistance with or without PTX treatment. Moreover, deletion of TUG1 in ovarian cancer cells decreased autophagosome formation and increased apoptosis as demonstrated by autophagy/cytotoxicity dual staining and Western blot assays. Furthermore, microRNA-29b-3p (miR-29b-3p) was found as the direct target of TUG1. Additionally, TUG1 could directly bind Ago2, a key protein of the RNA-induced silencing complex. CONCLUSION: Our findings suggest that TUG1, through targeting miR-29b-3p, induces autophagy and consequently results in PTX resistance in ovarian cancer.

Circular BANP, an upregulated circular RNA that modulates cell proliferation in colorectal cancer.

Circular RNAs (circRNAs) are recently identified as widespread and diverse endogenous noncoding RNAs that may harbor vital functions in human and animals. However, the role of circRNAs in the process of tumorigenesis and development of colorectal cancer (CRC) remains vague. Here we characterized the circRNA expression profile from three paired CRC cancerous and adjacent normal tissues by human circRNA array, and identified 136 significantly overexpressed circRNAs and 243 downregulated circRNAs in CRC cancerous tissues (>2-fold changes). We further validated one circRNA generated from Exon 5-11 of BANP gene, termed circ-BANP. In addition, RT-PCR result showed that circ-BANP was overexpressed in 35 CRC cancerous tissues. Knockdown of circ-BANP with siRNA significantly attenuate the proliferation of CRC cells. In summary, our findings demonstrated that dysregulated circ-BANP appears to have an important role in CRC cells and could serve as a prognostic and therapeutic marker for CRC.

RNAi-mediated downregulation of cyclin Y to attenuate human breast cancer cell growth.

Cyclin Y (CCNY) is a newly identified PFTK1 interacting protein and has been found to be associated with the proliferation and tumorigenesis of human non-small cell lung cancer. In the present study, we analyzed the expression levels of CCNY in 65 cases of breast cancer (BC) tissues and in four BC cell lines, BT-474, MDA-MB-231, T-47D and MCF-7. Lentivirus-mediated short hairpin RNA (shRNA) was employed to knock down CCNY expression in MCF-7 and MDA-MB-231 cells. The effects of CCNY depletion on cell growth were examined by MTT, colony formation and flow cytometry assays. The results showed that immunohistochemical expression of CCNY in tumor tissues is stronger than that in normal tissues. CCNY was also expressed in all four BC cells. The knockdown of CCNY resulted in a significant reduction in cell proliferation and colony formation ability. Cell cycle analysis showed that CCNY knockdown arrested MDA-MB­231 cells in the G0/G1 phase. Furthermore, depletion of CCNY inhibited BC cell growth via the activation of Bad and GSK3ß, as well as cleavages of PARP and caspase-3 in a p53-dependent manner. Therefore, we believe that CCNY has biological effect in BC development, and its inhibition via an RNA interference lentiviral system may provide a therapeutic option for BC.

Direct detection of circulating free DNA extracted from serum samples of breast cancer using locked nucleic acid molecular beacon.

As an emerging noninvasive blood biomarker, circulating free DNA (cfDNA) can be utilized to assess diagnosis, progression and evaluate prognosis of cancer. However, cfDNAs are not "naked", they can be part of complexes, or are bound to the surface of the cells via proteins, which make the detection more challenging. Here, a simple method for the detection of Ubiquitin-like with PHD and ring finger domains 1 (UHRF1) DNA exacted from serum of breast cancer (BC) has been developed using a novel locked nucleic acid molecular beacon (LNA-MB). In order to enhance the stability and detection efficiency of the probe in biofluids, we design a shared-stem molecular beacon containing a 27-mer loop and a 4-mer stem with DNA/LNA alternating bases. The fluorescence is released in the presence of target. The detection procedure is simple and can be completed within 1h. This method shows a sensitive response to UHRF1 DNA with a dynamic range of 3 orders of magnitude. The limit of detection is 11nM (S/N=3) with excellent selectivity. It can discriminate UHRF1 DNA from three-base mismatched DNA with a high specificity. More importantly, this method can distinguish the expression of serum UHRF1 DNA among 5 breast cancer patients and 5 healthy controls. The mentioned superiority may suggest that this assay can be served as a promising noninvasive detection tool for early BC diagnosis and monitoring.

Quantitative assessment of human serum transferrin receptor in breast cancer patients pre- and post-chemotherapy using peptide immunoaffinity enrichment coupled with targeted proteomics.

BACKGROUND: sTfR, a soluble form of transferrin receptor in serum, has been suggested as an indicator of bone marrow failure in breast cancer patients receiving chemotherapy. However, intensive chemotherapy could also cause a reduction of sTfR to a level below the LOQ of most assays. METHODS: An advanced liquid chromatography-tandem mass spectrometry (LC-MS/MS)-based targeted proteomics assay coupled with peptide immunoaffinity enrichment (SISCAPA) was developed and validated for monitoring sTfR. RESULTS: Tryptic peptide 681VEYHFLSPYVSPK693 was selected as a surrogate analyte for quantification. High-abundant proteins were first removed from serum, followed by SISCAPA that was effective in surrogate peptide enrichment and sensitivity enhancement. The resulting LOQ can achieve 100ng/ml (~10-fold increase). Then, sTfR levels in breast cancer patients pre- and post-chemotherapy, and healthy volunteers were accurately quantified as 1.77±0.53µg/ml, 0.98±0.26µg/ml and 1.66±0.50µg/ml, respectively, using a standard addition method. While there was no evidence for a difference between patients and healthy volunteers, differential levels of sTfR pre- and post-chemotherapy were obtained. Comparison between SISCAPA-targeted proteomics and ELISA indicated that the former approach provided a lower value of sTfR. CONCLUSIONS: SISCAPA-targeted proteomics may allow the quantification of low-abundant proteins in a more accurate manner.

Regulation of UHRF1 by microRNA-9 modulates colorectal cancer cell proliferation and apoptosis.

The UHRF1 protein is pivotal for DNA methylation and heterochromatin formation, leading to decreased expressions of tumor suppressor genes and contributing to tumorigenesis. However, the factors that modulate UHRF1 expression in colorectal cancer (CRC) remain unclear. Here we showed that, compared with corresponding normal tissues, UHRF1 was upregulated and microRNA-9 (miR-9) was downregulated in CRC tissues. The expression of UHRF1 was inversely correlated with overall survival rates of patients with CRC. Overexpression of miR-9 in CRC cell lines significantly attenuated CRC cell proliferation and promoted cell apoptosis. The expression of UHRF1 was markedly reduced in pre-miR-9 transfected CRC cells. Using luciferase reporter assay, we confirmed that miR-9 was a direct upstream regulator of UHRF1. Finally, analysis of miR-9 and UHRF1 levels in human CRC tissues revealed that expression of miR-9 was inversely correlated with UHRF1 expression. Collectively, our results offer in vitro validation of the concept that miR-9 could repress the expression of UHRF1, and function as a tumor-suppressive microRNA in CRC. It may serve as a prognostic and therapeutic marker for CRC.

Overexpression of metastasis-associated in colon cancer-1 associated with poor prognosis in patients with esophageal cancer.

Recent studies have shown that expression of metastasis-associated in colon cancer-1(MACC1) is observed in different types of cancer and plays an important role in tumor metastasis. However, the expression of MACC1 and its possible role in esophageal cancer remains unknown. In this study, we determined the expression of MACC1 in esophageal cancer by utilizing immunohistochemistry and analyzed the relationship between the expression and esophageal cancer prognosis. Immunohistochemistry results showed that 47 of 85 cancer lesions (55.2 %) were stained positive, and high expression of MACC1 was correlated with the node metastasis and TNM stage (P < 0.05). The Kaplan-Meier survival curve showed that patients with high MACC1 expression had significantly reduced overall 5-year survival rates (P = 0.004). Cox regression analysis revealed that high expression of MACC1 was associated with increased risk of death (hazard ratio [HR] =2.25) in patients with esophageal cancer. These findings suggested that high expression of MACC1 was correlated with progression and metastasis of esophageal cancer and might serve as a novel prognostic marker for patients with esophageal cancer.

Overexpressed ostepontin-c as a potential biomarker for esophageal squamous cell carcinoma.

BACKGROUND: The metastasis gene osteopontin (OPN) is subject to alternative splicing, which yields three messages, osteopontin-a, osteopontin-b and osteopontin-c. Osteopontin-c is selectively expressed in invasive, but not in noninvasive tumors. In the present study, we examined the expression of OPN-c in esophageal squamous cell carcinomas (ESCCs) and assessed its value as a diagnostic biomarker. METHODS: OPN-c expression was assessed by immunohistochemistry in 63 ESCC samples and correlated with clinicopathologic factors. Expression was also examined in peripheral blood mononuclear cells (PBMCs) from 120 ESCC patients and 30 healthy subjects. The role of OPN-c mRNA as a tumor marker was investigated by receiver operating characteristic curve (ROC) analysis. RESULTS: Immunohistochemistry showed that OPN-c was expressed in 30 of 63 cancer lesions (48%)and significantly associated with pathological T stage (P=0.038) and overall stage (P=0.023). Real time PCR showed that OPN-c mRNA was expressed at higher levels in the PBMCs of ESCC patients than in those of healthy subjects (P<0.0001) with a sensitivity as an ESCC biomarker of 86.7%. CONCLUSION: Our findings suggest that expression of OPN-c is significantly elevated in ESCCs and this upregulation could be a potential diagnostic marker.

Down-regulation of HLA-A mRNA in peripheral blood mononuclear cell of colorectal cancer.

PURPOSE: It has been demonstrated that the alteration of human leukocyte antigen (HLA) class I expression frequently occurs in colorectal tumor. Previous studies mainly focused on the expression of HLA-A in tumor cells. The expression of HLA-A in peripheral blood mononuclear cells (PBMC) was unknown. To develop a non-invasive diagnostic method for colorectal cancer (CRC), this work investigated the expression of HLA-A mRNA in PBMC in patients with CRC. METHODS: Real-time quantitative RT-PCR was used to study the expression of HLA-A mRNA in PBMC from 48 patients with colorectal cancer, 38 patients with benign colorectal lesions, 20 patients with rheumatoid arthritis, 20 patients with esophageal cancer and 40 healthy individuals. Protein chip was utilized to detect the levels of serum CEA, CA 19-9, and CA 242 in all the cases. Overall results from the two methods were compared. RESULTS: The relative expression of HLA-A mRNA in PBMC was 1.11 ± 0.45 in healthy group, 0.81 ± 0.42 in benign colorectal lesion group, and 0.39 ± 0.34 in cancer group, respectively. The diagnostic sensitivity of HLA-A mRNA, CEA, CA19-9, and CA242 was 81%, 59%, 61%, and 63%, and their diagnostic specificity was 75%, 64%, 52%, and 67%, respectively. CONCLUSIONS: The expression of HLA-A mRNA in PBMC from colorectal cancer group was significantly lower than those in both benign group and healthy group (P < 0.001). It could be potentially developed as a tumor assistant marker in future.

HLA Class I Expressions on Peripheral Blood Mononuclear Cells in Colorectal Cancer Patients.

OBJECTIVE: To investigate the expression change of human leukocyte antigen (HLA) class I on human peripheral blood mononuclear cells (PBMCs) at both mRNA and protein levels, and to evaluate its roles in the development of colorectal cancer (CRC). METHODS: In the present study, 50 patients with CRC, 35 patients with benign colorectal lesion and 42 healthy volunteers were enrolled. Expression levels of HLA class I mRNA and protein were determined using real-time quantitative reverse transcription PCR (RT-PCR) and flow cytometry analysis, respectively. RESULTS: The expression levels of HLA class I mRNA and proteins were not influenced by age and gender. The relative ratios of HLA class I mRNA were 0.99±0.27 in healthy controls, 0.76±0.19 in benign patients, and 0.48±0.21 in CRC patients. Mean fluorescence intensities of HLA class I were 145.58±38.14 in healthy controls, 102.05±35.98 in benign patients and 87.44±34.01 in CRC patients. HLA class I on PBMCs was significantly down-regulated at both mRNA and protein levels in patients with stage III and IV CRC. CRC patients with lymph node metastasis also showed a decreased HLA class I expression at protein level. CONCLUSION: HLA class I expressions on PBMCs are associated with staging of CRC and lymph node metastasis. Monitoring the expression of HLA class I on PBMCs may provide useful information for diagnosis and metastasis judgement of CRC.

Inhibition effect of siRNA-downregulated UHRF1 on breast cancer growth.

The UHRF1 gene plays important roles in both cell proliferation through its NIRF_N domains, a PHD domain, an SRA domain, and a RING domain, and multidrug resistance in breast cancer treatment. In this work, a short-hairpin RNA (shRNA) lentiviral system was introduced in two human breast cancer cell lines (MDA-MB-231 and MCF-7) to downregulate the expression of UHRF1 and study the specific inhibition of UHRF1 in breast cancer growth. The effect of UHRF1-shRNA on breast cancer cell proliferation was examined using methylthiazoletetrazolium, bromodeoxyuridine, and colony formation assays. The proliferative potential of the UHRF1-shRNA-treated cells showed a remarkable decrease. Moreover, the downregulation of UHRF1 in both breast cancer cell lines significantly inhibited the colony formation capacity. Results suggested that the inhibition of UHRF1 via an RNA interference lentiviral system may provide an effective way for breast cancer therapy.

Expression and prognostic value of PRL-3 in human intrahepatic cholangiocarcinoma.

Phosphatase of regenerating liver (PRL)-3 is involved in the metastasis of various tumors, but the expression of PRL-3 and its possible role in primary intrahepatic cholangiocarcinoma (ICC) has not been reported yet. In this study, we assessed the expression levels of PRL-3 by immunohistochemistry in 102 primary ICC samples, 62 matched lymph node metastases (LNM) and 102 adjacent normal liver tissues. Then we investigated the relationship between PRL-3 expression and clinicopathologic factors. Survival analysis was performed to determine the prognostic significance of PRL-3 expression in ICC. Immunochemistry results suggested PRL-3 expression was negative or weak in non-neoplastic intrahepatic bile ducts of adjacent liver tissue. In primary lesion and LNM high PRL-3 expression was frequently detected. Furthermore, the rate of high PRL-3 expression in LNM was higher than that in primary lesion (80.6% vs. 47.1%, P < 0.05). High expression of PRL-3 in primary tumors was significantly associated with TNM (P < 0.001), T stage (P < 0.001), vascular invasion (P = 0.002), and LNM (P < 0.001). Survival analysis results with Kaplan-Meier method and Cox proportional hazard model indicated high expression of PRL-3 was correlated with decreased overall survival and was an independent prognostic marker of overall survival. Thus, our results suggested high expression of PRL-3 was correlated with progression and metastasis of ICC and indicated negative prognostic impact. PRL-3 might serve as a novel prognostic marker for patients with ICC.

Qa-2 mRNA in peripheral blood mononuclear cells: a potential marker of acute allograft rejection.

BACKGROUND: HLA-G displays an immunotolerogenic role and its expressions in grafts/sera are related to allograft acceptance. However, it is still unclear how its transcription level in peripheral blood mononuclear cells (PBMCs) changes during allograft rejection. The aim of the present study was to detect the expression changes of the murine homolog of HLA-G, Qa-2, serially during the whole process of graft rejection with mouse skin transplantation models and to investigate their relationship with the pathological grade of graft rejection and immunosuppressive therapy. MATERIAL/METHODS: Full-thickness skin derived from donor mice (C57BL/6j for syngeneic groups and Balb/c for allogeneic groups) was transplanted onto the dorsal thorax of recipient mice (C57BL/6j). Pathological allograft changes were classified into four grades. Qa-2 mRNA expression of PBMCs was detected serially by real-time RT-PCR. RESULTS: Qa-2 mRNA did not show any obvious change in the syngeneic recipients without immunosuppressive treatment, inferring that surgical stress might not influence Qa-2 expression. A significant increase in Qa-2 mRNA was observed in the immunosuppressant-treated recipients, suggesting that the increased Qa-2 mRNA level might be attributed to the immunosuppressive treatment. During allograft rejection, the Qa-2 mRNA level decreased significantly, especially with immunosuppressive treatment, and the decreased expression was detected when the allograft presented grade 2-3 pathological rejection, much earlier than when gross rejection was observed. CONCLUSIONS: These results suggest that decreased expression of Qa-2 mRNA in PBMCs may be a potential marker for predicting acute allograft rejection as well as a tool to monitor the effects of immunosuppressive treatment.