Assessment of mucin-related gene alterations following treatment with rebamipide ophthalmic suspension in Sjögren's syndrome-associated dry eyes.

Ocular surface mucins are thought to play vital roles in maintaining the homeostasis of the pre-ocular surface tear film. We performed ocular surface tests with impression cytology to assess the expression levels of mucin-related genes on the ocular surface in healthy eyes. In addition, we investigated alterations in mucin-related gene expression secondary to treatment with rebamipide ophthalmic suspension in patients with Sjögren's syndrome-associated dry eyes (SS-DE). Thirty-three healthy individuals (control group) and 13 patients from our hospital with SS-DE were enrolled. Impression cytology was performed using Schirmer's test paper for RNA sampling. The mRNA levels of SAM-pointed domain-containing ETS-like factor (SPDEF), mucin 5AC (MUC5AC), and mucin 16 (MUC16) were determined using a real-time reverse transcription-polymerase chain reaction. The ocular surface test was performed once for the control group, and at baseline as well as 2, 4, 8, and 12 weeks after treatment in the Sjögren's syndrome-associated dry eyes group. mRNA levels of SPDEF, MUC5AC, and MUC16 were not significantly different between the control and SS-DE groups before rebamipide ophthalmic suspension treatment. SPDEF mRNA levels in control subjects were significantly correlated with levels of MUC5AC. Among SS-DE patients, SPDEF mRNA levels were significantly increased at 2, 4, and 8 weeks after treatment compared with baseline levels. MUC16 mRNA levels were significantly decreased from baseline levels at 4 and 8 weeks post-treatment. Ocular surface test using impression cytology is a clinically useful tool for assessing mucous conditions on the ocular surface and can be used to determine the effects of instillation treatment with eye drops that affect mucin production at the ocular surface.

ERO1L promotes IL6/sIL6R signaling and regulates MUC16 expression to promote CA125 secretion and the metastasis of lung cancer cells.

The abnormal secretion of CA125, a classic tumor marker, is usually related to a poor prognosis in various tumors. Thus, this study aimed to explore the potential mechanisms that promote CA125 secretion in lung cancer. By querying the database, the gene endoplasmic reticulum oxidoreductase 1L (ERO1L) was identified and chosen as the research subject. The antibody chips were used to screen the lung cancer cell supernatant and found that the most obvious secreted protein was CA125. ERO1L was found to promote the secretion of IL6R by affecting the formation of disulfide bonds. IL6R bound to IL6 and triggered the activation of the NF-κB signaling pathway. Then, NF-κB bound to the promoter of MUC16, resulting in overexpression of MUC16. The extracellular segment of MUC16 was cleaved to form CA125, while the C terminus of MUC16 promoted the EMT phenotype and the release of IL6, forming a positive feedback pathway. In conclusion, ERO1L might affect the secretion of CA125 through the IL6 signaling pathway and form a positive feedback loop to further promote the development of lung cancer. This might expand the application scope of CA125 in lung cancer.

Postoperative expressions of TRACP5b and CA125 in patients with breast cancer and their values for monitoring bone metastasis.

PURPOSE: To explore the diagnostic values of serum tartrate-resistant acid phosphatase 5b (TRACP5b) and serum carbohydrate antigen 125 (CA125) for bone metastasis of breast cancer. METHODS: 118 patients pathologically diagnosed with breast cancer in the second People's Hospital of Lianyungang from September 2014 to June 2017 were selected. Among them, 60 patients who were confirmed with bone metastasis by whole-body bone imaging combined with clinical manifestations and other imaging methods were included in a bone metastasis group, and 58 patients who were confirmed without bone metastasis were included in a non-bone metastasis group. Another 61 patients who were pathologically confirmed with benign breast lesion formed a benign lesion group. Enzyme-linked immunosorbent assay (ELISA) was used to detect TRACP5b level and electrochemiluminescence (ECL) was used to detect CA125 level. RESULTS: The expression levels of TRACP5b and CA125 in the bone metastasis group were significantly higher than those in the non-bone metastasis and benign lesion groups (p<0.05), and the expression levels in the non-bone metastasis group were higher than those in the benign lesion group (p<0.05). In bone metastasis of breast cancer, the expression level of TRACP5b was correlated with the number of tumor nodules, lymph node metastasis, tumor local infiltration and TNM staging (p<0.05), while the expression level of CA125 was correlated with the number of tumor nodules, lymph node metastasis and TNM staging (p<0.05). Logistic regression analysis showed that TNM staging, estrogen receptor (ER), TRACP5b, and CA125 were risk factors for bone metastasis of breast cancer patients. CONCLUSION: In conclusion, TRACP5b and CA125 may be involved in the occurrence and progression of bone metastasis of breast cancer. Detection of TRACP5b and CA125 has good sensitivity and specificity in diagnosing bone metastasis of breast cancer, so TRACP5b and CA125 may become new biomarkers for diagnosing the disease.

Recurrent endometriosis: a battle against an unknown enemy.

Recurrence of endometriosis after conservative surgery is not an uncommon finding. There is no uniformity, however, on what the term 'recurrence' means. Recurrence is variously defined in the literature as the relapse of pain, clinical or instrumental detection of an endometriotic lesion, repeat rise in CA 125 levels, or evidence of recurrence found during repeat surgery. Consequently, the reported recurrence rate varies widely (0-89%) in the different series, depending on its definition and the type of study performed. As endometriosis recurrence seems to be an indeterminate enemy, we set out to examine exactly what we were fighting in our everyday battle. In this narrative review, we aimed to seek an answer to questions related to endometriosis recurrence, some of which are often asked by our patients.

A 2-Protein Signature Predicting Clinical Outcome in High-Grade Serous Ovarian Cancer.

OBJECTIVE: High-grade serous ovarian cancer (HGSOC) accounts for approximately 70% deaths in ovarian cancer. The overall survival (OS) of HGSOC is poor and still remains a clinical challenge. High-grade serous ovarian cancer can be divided into 4 molecular subtypes. The prognosis of different molecular subtypes is still unclear. We aimed to investigate the prognostic values of immunohistochemistry-based different molecular subtypes in patients with HGSOC. METHODS: We analyzed the protein expression of representative biomarkers (CXCL11, HMGA2, and MUC16) of 3 different molecular subtypes in 110 formalin-fixed, paraffin-embedded HGSOC by tissue microarrays. RESULTS: High CXCL11 expression predicted worse OS, not disease-free survival (DFS; P = 0.028 for OS, P = 0.191 for DFS). High HMGA2 expression predicted worse OS and DFS (P = 0.037 for OS, P = 0.021 for DFS). MUC16 expression was not associated with OS or DFS (P = 0.919 for OS, P = 0.517 for DFS). Multivariate regression analysis showed that CXCL11 combined with HMGA2 signature was an independent predictor for OS and DFS in patients with HGSOC. CONCLUSIONS: CXCL11 combined with HMGA2 signature was a clinically applicable prognostic model that could precisely predict an HGSOC patient's OS and tumor recurrence. This model could serve as an important tool for risk assessment of HGSOC prognosis.

Oncogenic KRAS Targets MUC16/CA125 in Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is a devastating disease with the 5-year survival rate less than 6%. Previous results indicated that serum levels of CA125 (encoded by MUC16) could be used to predict which groups of pancreatic cancer patients may benefit from surgery. However, the underlying mechanism remains elusive. Herein, using the Cancer Genome Atlas and clinicopathologic data obtained from our center, we demonstrate that high CA125 serum levels and expression levels of MUC16 are predictive of poor prognosis. MUC16 is also validated as a downstream target of KRAS, and their expression strongly correlated with each other in vitro and in vivo Mechanistically, the KRAS/ERK axis induced upregulation of MUC16 and shedding of CA125 via its effector c-Myc in SW1990 and PANC-1 pancreatic cancer cells. Notably, proto-oncogene c-Myc could bind to the promoter of MUC16 and transcriptionally activate its expression. Taken together, these data establish CA125 as a prognostic marker for pancreatic cancer, and mechanistic studies uncovered the KRAS/c-Myc axis as a driving factor for upregulation of MUC16. IMPLICATIONS: The current study uncovers the contribution of oncogenic KRAS to serum marker CA125 production through a mechanism that involves the ERK/c-Myc axis. Mol Cancer Res; 15(2); 201-12. ©2016 AACR.

PPARγ Modulation of Cytokine-Stimulated MUC16 (CA125) Expression in Breast and Ovarian Cancer-Derived Cells.

CA125 is serum tumor marker consisting of an epitope carried by a portion of the extremely large (>3 MDa), heavily glycosylated cell surface transmembrane mucin, MUC16. In malignancies, membrane bound mucins lose their polarized distribution, become aberrantly over-expressed and protect tumor cells from the actions of chemotherapeutic agents as well as the immune system. Previously, we described stimulation of MUC16 expression by the proinflammatory cytokines, tumor necrosis factor α (TNFα) and interferon γ (IFNγ), in breast and ovarian cancer cells and tissues. Herein, we show that PPARγ modulates cytokine-stimulated MUC16 in a complex manner: at low concentrations (<10 µM) rosiglitazone further potentiates cytokine-driven MUC16 expression while at high concentrations (>20 µM) rosiglitazone antagonizes cytokine stimulation. Rosiglitazone actions were fully reversible by the PPARγ antagonist, GW9662. Furthermore, siRNA-mediated PPARγ knockdown also prevented a large portion of high dose rosiglitazone suppression of MUC16 expression indicating that rosiglitazone inhibition is largely PPARγ-dependent. Cytokines greatly (>75%) suppressed PPARγ expression. Conversely, PPARγ activation by rosiglitazone at either low or high concentrations greatly (>75%) suppressed NFκB/p65 expression. NFκB/p65 expression was largely preserved in the presence of cytokines at low, but not high, rosiglitazone concentrations accounting for the different concentration dependent effects on MUC16 expression. Collectively, these studies demonstrate that PPARγ is an important modulator of MUC16 expression. The ability to deliver high doses of PPARγ agonists to MUC16-expressing tumors offers an avenue to reduce expression of this protective glycoprotein and increase tumor sensitivity to killing by chemotherapeutic drugs and the immune system. J. Cell. Biochem. 118: 163-171, 2017. © 2016 Wiley Periodicals, Inc.

Placental type alkaline phosphatase tissue expression in ovarian serous carcinoma.

OBJECTIVE: To analyze the expression profile of placental type alkaline phosphatase (PLAP), cancer antigen 125 (CA125), and human epididymis protein 4 (HE4) in serous ovarian cancer and to correlate their expression with the tumor aggressiveness and progression. METHODS: Retrospective study considering a tissue microarray of 82 women affected by ovarian serous cancer. Protein expression was assessed by immunohistochemistry on ovarian serous cancer tissue samples. Immunohistochemical staining was semiquantitatively evaluated as H-score. RESULTS: Median H-score values were lower for PLAP, 1 (IQR 0-4) than CA125, 10 (IQR 6-12) or HE4, 8 (IQR 5-12). Even if PLAP was less expressed in the cells of serous ovarian cancer than CA125 or HE4 it was relatively more expressed in the fourth quartile of its H-score distribution among cases with low CA125 or HE4 expression. Furthermore, PLAP and HE4 high expression resulted to be significantly correlated with a better prognosis. CONCLUSIONS: PLAP could be an additional marker for early detection of serous ovarian carcinoma, together with the established CA125 and HE4. In addition, PLAP expression is correlated with prognosis, giving, in this way, an additional tool for improving treatment approach.

Differential effect of rebamipide on transmembrane mucin biosynthesis in stratified ocular surface epithelial cells.

Mucins are a group of highly glycosylated glycoproteins responsible for the protection of wet-surfaced epithelia. Recent data indicate that transmembrane mucins differ in their contribution to the protective function of the ocular surface, with MUC16 being the most effective barrier on the apical surface glycocalyx. Here, we investigated the role of the mucoprotective drug rebamipide in the regulation of transmembrane mucin biosynthesis using stratified cultures of human corneal and conjunctival epithelial cells. We find that the addition of rebamipide to corneal, but not conjunctival, epithelial cells increased MUC16 protein biosynthesis. Rebamipide did not affect the levels of MUC1, 4 and 20 compared to control. In these experiments, rebamipide had no effect on the expression levels of Notch intracellular domains, suggesting that the rebamipide-induced increase in MUC16 biosynthesis in differentiated corneal cultures is not regulated by Notch signaling. Overall these findings indicate that rebamipide induces the differential upregulation of MUC16 in stratified cultures of human corneal epithelial cells, which may have implications to the proper restoration of barrier function in ocular surface disease.

Mesothelial cells interact with tumor cells for the formation of ovarian cancer multicellular spheroids in peritoneal effusions.

Epithelial ovarian cancer (EOC) dissemination is primarily mediated by the shedding of tumor cells from the primary site into ascites where they form multicellular spheroids that rapidly lead to peritoneal carcinomatosis. While the clinical importance and fundamental role of multicellular spheroids in EOC is increasingly appreciated, the mechanisms that regulate their formation and dictate their cellular composition remain poorly characterized. To investigate these important questions, we characterized spheroids isolated from ascites of women with EOC. We found that in these spheroids, a core of mesothelial cells was encased in a shell of tumor cells. Analysis further revealed that EOC spheroids are dynamic structures of proliferating, non-proliferating and hypoxic regions. To recapitulate these in vivo findings, we developed a three-dimensional co-culture model of primary EOC and mesothelial cells. Our analysis indicated that, compared to the OVCAR3 cell line, primary EOC cells isolated from ascites as well as mesothelial cells formed compact spheroids. Analysis of heterotypic spheroid microarchitecture revealed a structure that grossly resembles the structure of spheroids isolated from ascites. Cells that formed compact spheroids had elevated expression of ß1 integrin and low expression of E-cadherin. Addition of ß1 integrin blocking antibody or siRNA-mediated downregulation of ß1 integrin resulted in reduced tightness of the spheroids. Interestingly, the loss of MUC16 and E-cadherin expression resulted in the formation of more compact spheroids. Therefore, our findings support the heterotypic nature of spheroids from malignant EOC ascites. In addition, our data describe an unusual link between E-cadherin expression and less compact spheroids. Our data also emphasize the role of MUC16 and ß1 integrin in EOC spheroid formation.

Upregulation of centrosomal protein 55 is associated with unfavorable prognosis and tumor invasion in epithelial ovarian carcinoma.

Centrosomal protein 55 (CEP55) is a cell cycle regulator implicated in development of certain cancers. However, characteristics of CEP55 expression and its clinical/prognostic significance are unclear in human epithelial ovarian carcinoma (EOC). Therefore, we investigated the expression and clinicopathological significance of CEP55 in patients with EOC and its role in regulating invasion and metastasis of ovarian cell lines. CEP55 mRNA and protein expression levels were detected by quantitative real-time PCR (qRT-PCR), Western blotting, and immunohistochemistry (IHC). Potential associations of CEP55 expression scores with clinical parameters and patient survival were evaluated. CEP55 function was investigated further using RNA interference, wound healing assay, transwell assay, immunofluorescence analysis, qRT-PCR, and Western blotting. CEP55 was significantly upregulated in ovarian cancer cell lines and lesions compared with normal cells and adjacent noncancerous ovarian tissues. In the 213 EOC samples, CEP55 protein levels were positively correlated with clinical stage (P < 0.001), lymph node metastasis (P < 0.001), intraperitoneal metastasis (P < 0.001), tumor recurrence (P < 0.001), differentiation grade (P < 0.001), residual tumor size (P < 0.001), ascites see tumor cells (P = 0.020), and serum CA153 level (P < 0.001). Moreover, patients with aberrant CEP55 protein expression showed tendencies to receive neoadjuvant chemotherapy (P < 0.001) and cytoreductive surgery (P = 0.020). By contrast, no significant correlation was detected between the protein levels and patient age, histological type, or serum CA125, CA199, CA724, NSE, CEA, and ß-HCG levels. Patients with high CEP55 protein expression had shorter overall survival and disease-free survival compared with those with low CEP55 expression. Multivariate analysis implicated CEP55 as an independent prognostic indicator for EOC patients. Additionally, downregulation of CEP55 in ovarian cancer cells remarkably inhibited cellular motility and invasion. Aberrant CEP55 expression may predict unfavorable clinical outcomes in EOC patients and play an important role in regulating invasion in ovarian cancer cells. Thus, CEP55 may serve as a prognostic marker and therapeutic target for EOC.

The role of HE4 in endometrial cancer recurrence: how to choose the optimal follow-up program.

The aim of this study was to evaluate for the first time in the literature the role of HE4, at primary diagnosis, compared to CA125 as an indicator of endometrial cancer (EC) recurrence. Our study is a retrospective analysis of 252 EC patients treated, between January 2009 and July 2013, at the Division of Gynaecologic Oncology of Campus Bio-Medico University of Rome. Thirty-seven patients experienced recurrence. Median follow-up was 38 months. HE4 and CA125 levels were analyzed at primary diagnosis, during follow-up and either after histological or radiological confirmation of recurrent disease or at last registered visit, when patients returned to our Department with no evidence of recurrent disease. A statistically significant difference was observed between HE4 values at primary diagnosis and at recurrence, respectively, comparing recurrent and non-recurrent patients (p < 0.05), while CA125 values resulted not statistically significant (p = 0.08) at each time point. Considering the poor specificity of HE4 at threshold of 70 pmol/L at primary diagnosis, in our cohort of patients, we found out that HE4 cut-off of 201.3 pmol/L is able to correctly classify patients at high or low risk of EC recurrence, with a sensitivity of 80 % and a specificity of 91 % (PPV = 90.3 % and NPV = 90.8 %). In particular, HE4 performance improves in cases of endometrioid histotype. HE4 levels at primary diagnosis correlate with an increased risk of EC recurrence, particularly in cases of endometrioid histotype, and they may help to recognize patients who may need a more intensive follow-up.

Analysis of laparoscopy on endometriosis patients with high expression of CA125.

OBJECTIVE: To analyze the clinical effect of laparoscopy on patients with endometriosis and a high expression of CA125. PATIENTS AND METHODS: One hundred cases of endometriosis treated in our hospital from May 2012 to August 2014 were selected as research subjects, after approval from the hospital and the informed consent of patients. The subjects were randomly divided into a control group and a study group with 50 cases in each group. The control group underwent the traditional surgery, while the research group underwent laparoscopy. The clinical curative effect were compared between the two groups of patients. RESULTS: The research group's postoperative 6h, 12h, 24h CA125 levels were compared to those of the control group. There was statistically significant difference between groups (p < 0.05). Surgery and hospitalization time, intraoperative bleeding volume, and and incision length index of the two groups of patients were compared. There were statistically significant differences between the two groups (p < 0.05). CONCLUSIONS: Laparoscopic surgery in endometriosis patients with high expression of CA125 is curative, and the patients'recovery after surgery is promising.

Relationship of tumor marker CA125 and ovarian tumor stem cells: preliminary identification.

PURPOSE: The purpose of this study is to identify a prospective association between CA125 and tumorigenic ovarian cancer cells, using the new method of orthotopic transplantation (1). METHOD: After making the surgical ovarian cancer specimen into cell suspension, we separated the tumorigenic cells from the nontumorigenic cancer cells based on cell surface marker (cancer antigen CA125 and lineage markers) expression. We developed a SCID mice model in which the CA125+/ lineage- and CA125-/ lineage- cells were injected into ovarian parenchyma by use of a microinjector. As a measure of effectiveness of tumor-forming, tumor weight, abdominal distension, ascites volume and activity, subcutaneous fat were determined or observed. Immunohistochemistry was done to determine tumor cell markers. RESULTS: We found that the cells of CA125+/ lineage- were able to form new tumors; whereas, an equal quantity of CA125-/lineage- cells failed to form any tumors. The new generated tumor contained additional CA125-/lineage- tumorigenic cells as well as the phenotypically diverse population of nontumorigenic cells. Quantities were judged to be significantly different P < 0.0001. CONCLUSION: CA125+/ lineage- cells, which may be ovarian cancer stem cells, were the source for tumor recurrence. The strategies designed to target this cell population may lead to more effective therapies.

Effects of the loss of conjunctival Muc16 on corneal epithelium and stroma in mice.

PURPOSE: To examine the role of conjunctival Muc16 in the homeostasis of the ocular surface epithelium and stroma using Muc16-null knockout (KO) mice. METHODS: We used KO mice (n = 58) and C57/BL6 (WT) mice (n = 58). Histology and immunohistochemistry were employed to analyze the phenotypes in the ocular surface epithelium. The expression of phospho-Stat3, AP-1 components, interleukin 6 (IL-6), and tumor necrosis factor-α (TNFα) in the cornea and conjunctiva was examined. The shape of the nuclei of corneal epithelial cells was examined to evaluate intraepithelial cell differentiation. Epithelial cell proliferation was studied using bromo-deoxyuridine labeling. Finally, the wound healing of a round defect (2-mm diameter) in the corneal epithelium was measured. The keratocyte phenotype and macrophage invasion in the stroma were evaluated after epithelial repair. RESULTS: The loss of Muc16 activated Stat3 signal, affected JunB signal, and upregulated the expression of IL-6 in the conjunctiva. Basal-like cells were observed in the suprabasal layer of the corneal epithelium with an increase in proliferation. The loss of Muc16 accelerated the wound healing of the corneal epithelium. The incidence of myofibroblast appearance and macrophage invasion were more marked in KO stroma than in WT stroma after epithelial repair. CONCLUSIONS: The loss of Muc16 in the conjunctiva affected the homeostasis of the corneal epithelium and stroma. The mechanism might include the upregulation of the inflammatory signaling cascade (i.e., Stat3 signal, and IL-6 expression in the KO conjunctiva). Current data provides insight into the research of the pathophysiology of dry eye syndrome.

Is tissue CA125 expression in epithelial ovarian adenocarcinoma heterogenic?

To evaluate if heterogeneity of tissue cancer antigen 125 (CA125) expression is present in epithelial serous adenocarcinomas. Furthermore, to investigate whether there is a correlation between levels of CA125 tissue expression, serum level of CA125, stage, and grade. A total of 10 patients diagnosed with serous ovarian adenocarcinomas were included. Preoperative blood samples were collected to determine serum CA125 levels. Tumor tissue from primary surgery was collected and processed for immunohistochemical analyses. CA125 was expressed in varying degrees in tumor tissues from all patients. Mean tissue CA125 expression for each patient ranged from 36% to 98%. Intrapatient variations in tissue expression ranged from 10% to 90% point. No significant correlations between levels of CA125 tissue expression, serum level of CA125, stage, and grade were found. We found that the tissue expression of CA125 is heterogenic. Although most patients had a high mean expression, it covers a large intrapatient variation in expression. This suggests that if using CA125 as a tissue marker and anti-CA125 (oregovomab) as immunotherapy treatment in future studies, it will be necessary to take heterogeneity into consideration and examine a larger number of biopsies. Therefore, the study demonstrates the need for heterogeneity studies in future translational research.

Hematopoietic cytokines as tumor markers in gynecological malignancies. A multivariate analysis in epithelial ovarian cancer patients.

We investigated plasma levels of selected hematopoietic cytokines: stem cell factor (SCF), granulocyte-macrophage colony-stimulating factor (GM-CSF), granulocyte colony-stimulating factor (G-CSF), and macrophage colony-stimulating factor (M-CSF) and the tumor marker cancer antigen (CA 125) in epithelial ovarian cancer patients as compared with control groups: benign ovarian tumor patients (cysts) and healthy subjects. Cytokine levels were determined by enzyme-linked immunosorbent assay, CA 125 - using the chemiluminescent microparticle immunoassay method. Our results have demonstrated significant differences in the concentrations of M-CSF, G-CSF, SCF (with the exception of GM-CSF), and CA 125 between the groups of ovarian cancer patients, cysts patients, and the healthy controls. When compared with CA 125, M-CSF has equal or higher values of diagnostic sensitivity and specificity. The M-CSF area under the receiver-operating characteristic curve (AUC) was the largest from all the cytokines tested and slightly lower than the AUC of CA 125. These findings suggest the usefulness of M-CSF in diagnosing ovarian cancer, especially when discriminating between cancer and non-carcinoma lesions.

[Expression and purification of mucin 16 and preparation and characterization of anti-mucin 16 monoclonal antibody].

AIM: To generate monoclonal antibodies (mAbs) against mucin 16 using purified recombinant protein of human mucin 16 N terminus with His tag (His-mucin 16N) as the antigen. METHODS: Mucin 16 N terminus was cloned into a prokaryotic expression vector pET-32. His-mucin 16N was then expressed in E.coli and purified by the affinity chromotography. Cell fusion was performed after the BALB/c mice were immunized with the purified His-mucin 16N protein. We screened hybridoma cell strains producing mAbs against mucin 16. The specificity and titer of the antibodies were characterized with ELISA, Western blotting, immunofluorescent and immunohistochemical staining. RESULTS: The recombinant protein of His-mucin 16N was expressed and purified. A few hybridoma cell strains which could secrete specific mAbs against mucin 16 were obtained, and one anti-mucin 16 mAb with good specificity and high titer was selected and purified. The isotype of this anti-mucin 16 mAb was determined as IgG1, which indicated that this anti-mucin 16 mAb could be used for ELISA, Western blotting, immunofluorescent and immunohistochemical staining. The endogenous expression of mucin 16 in various cancer cell lines or tissues was also examined with this anti-mucin 16 antibody by Western blotting and other immunoassays. CONCLUSION: The recombinant protein of His-mucin 16N was expressed and purified successfully, with which we prepared anti-mucin 16 mAb with good specificity and high titer.

New mechanism of elevated CA125 in heart failure: the mechanical stress and inflammatory stimuli initiate CA125 synthesis.

Carbohydrate antigen 125 has been known as a membrane-associated mucin. It has found application as a tumor marker that may be elevated in the blood of some patients with ovarian cancer, or other benign conditions. Recently, increased serum carbohydrate antigen 125 levels have been documented in patients with heart failure. However, little is known about the pathophysiologic mechanism. It has been found that carbohydrate antigen 125 will be shed from surfaces of mesothelial cells in response to mechanical stress such as fluid overload, and inflammatory stimuli. High carbohydrate antigen 125 levels are closely related to the presence of serosal fluid and positively correlated with serum TNF-α, IL-6 and IL-10 levels in heart failure patients. This leads to proposed hypothesis that the mechanical stress and inflammatory stimuli both initiate carbohydrate antigen 125 synthesis. This finding suggests that carbohydrate antigen 125 might be a promising biomarker to evaluate the risk stratification of heart failure and monitor the process of therapy.

Mucin 16 (cancer antigen 125) expression in human tissues and cell lines and correlation with clinical outcome in adenocarcinomas of the pancreas, esophagus, stomach, and colon.

Mucin 16 (cancer antigen 125) is a cell surface glycoprotein that plays a role in promoting cancer cell growth in ovarian cancer. The aims of this study were to examine mucin 16 expression in a large number of digestive tract adenocarcinomas and precursors and to determine whether mucin 16 up-regulation is correlated with patient outcome. Tissue microarrays were constructed using surgical resection tissues and included pancreatic (115 normal, 29 precursors, 200 pancreatic ductal adenocarcinomas), esophageal (86 normal, 104 precursors, 95 esophageal adenocarcinomas, 35 lymph node metastases), gastric (211 normal, 8 precursors, 119 gastric adenocarcinomas, 62 lymph node metastases), and colorectal (34 normal, 17 precursors, 39 colorectal adenocarcinomas) tissues. Mucin 16 was detected in 81.5%, 69.9%, 41.2%, and 64.1% of the pancreatic ductal adenocarcinomas, esophageal adenocarcinomas, gastric adenocarcinomas, and colorectal adenocarcinomas, respectively. Mucin 16 was seen in a subset of the precursors. On multivariate analysis, moderate/diffuse mucin 16 in pancreatic ductal adenocarcinomas was strongly associated with poor survival (P < .001), independent of other prognosis predictors. A similar trend was observed for esophageal adenocarcinomas (P = .160) and gastric adenocarcinomas (P = .080). Focal mucin 16 in colorectal adenocarcinomas was significantly correlated (P = .044) with a better patient outcome, when compared with mucin 16-negative cases. Using Western blot analysis, we found mucin 16 expression in 3 of 6 pancreatic ductal adenocarcinoma and 1 of 2 esophageal adenocarcinoma cell lines. We conclude that most of the digestive tract adenocarcinomas and a subset of their precursors express mucin 16. Mucin 16 expression is an independent predictor of poor outcome in pancreatic ductal adenocarcinomas and potentially in esophageal adenocarcinomas and gastric adenocarcinomas. We propose that mucin 16 may function as a prognostic marker and therapeutic target in the future.