The clinical significance of DJ-1 and HE4 in patients with endometrial cancer.

BACKGROUND: The non-invasive diagnostic approach for early detection of endometrial cancer (EC) remains limited. To date, human epididymis protein 4 (HE4) has been intensively studied but its diagnostic is controversial in EC. DJ-1 is an oncoprotein secreted by cancer cells, recently identified as a potential diagnostic biomarker for breast cancer, melanoma, and pancreatic cancer. The aim of this study was to compare the diagnostic performances of DJ-1 and HE4 measured in EC patients and healthy controls (HC). METHODS: Forty-five patients (63.9±12.0 years) with EC and 29 (63.2±13.3 years) HC were enrolled. Serum concentrations of DJ-1 and HE4 were measured using ELISA kits developed by R&D (Minneapolis, USA) and Fujirebio Diagnostic (Malvern, PA, USA), respectively. Differences between EC patients and HC were assessed by Mann-Whitney test and associations were tested by Spearman's correlation. The diagnostic performance was assessed using receiver operating characteristics (ROC) curves analysis. RESULTS: Serum DJ-1 concentrations were found to be higher in EC patients than in HC (9533.6 vs 1988.5 pg/mL; P<.0001). The area under the ROC curve (ROC-AUC) was 0.95 (P<.0001). At the cut-off of 3654 pg/mL, the sensitivity and specificity were 0.89 and 0.90, respectively. HE4 serum levels were higher in EC patients than in HC (75.3 vs 56.2 pmol/L; P=.019), with an AUC of 0.66 (P=.020). The AUC obtained by the combination of the two markers resulted 0.96 (P<.0001). CONCLUSION: These results suggest that increased serum DJ-1 levels are associated with EC and that this biomarker may be potentially useful for diagnosing EC.

Plasma Leptin in Patients at Intermediate to High Cardiovascular Risk With and Without Type 2 Diabetes Mellitus.

BACKGROUND: A number of clinical studies have demonstrated that leptin concentrations are related to the metabolic disturbances that constitute the metabolic syndrome (MetS) and to diabetes mellitus (DM). AIM: To investigate possible determinants of leptin concentrations in a sample of patients at high cardiovascular (CV) risk carrying two or more features of the MetS and to investigate if any difference exist between at risk patients with or without DM. METHODS: Serum leptin concentrations were measured in 60 consecutive male patients affected by at least two CV risk factors which belong to the National Cholesterol Education Program/Adult Treatment Panel III (NCEP/ATP III) definition of MetS: 30 patients affected by type 2 DM (T2DM) and 30 nondiabetic patients (non-T2DM). Nineteen healthy subjects were included in the study as a control group (HC). RESULTS: Leptin was significantly higher in patients carrying two or more features of the MetS compared with HC (P = 0.02). Stratifying MetS patients for DM, we found that leptin level was higher in non-T2DM patients (7.8 ng/ml), intermediate in T2DM (6.2 ng/ml), and lower in HC (4.6 ng/ml). In MetS patients, a positive correlation was found between leptin and waist, triglycerides, and number of MetS criteria. After stratification for T2DM, the correlations were still significant in the non-T2DM but not in the T2DM group. CONCLUSIONS: In our sample of moderate-to-high-risk patients, leptin level is positively associated with waist circumference and triglycerides but only in non-T2DM patients. Our data suggest that diabetic subjects could modulate leptin production in a different way compared with patients carrying other MetS-related anomalies.

The ROMA (Risk of Ovarian Malignancy Algorithm) for estimating the risk of epithelial ovarian cancer in women presenting with pelvic mass: is it really useful?

BACKGROUND: The study is aimed at evaluating the performance of the predictive model ROMA (Risk of Ovarian Malignancy Algorithm), which utilizes the combination of human epididymis protein 4 (HE4) and CA125 values to assess the risk of epithelial ovarian cancer (EOC) in women with a pelvic mass. METHODS: One hundred and four women diagnosed with a pelvic mass (55 EOC and 49 benign cases) and scheduled to have surgery were enrolled, along with 49 healthy females. Preoperative serum concentrations of HE4 and CA125 were measured. Separate logistic regression algorithms ROMA for pre-menopausal and post-menopausal women were used to categorize patients into low- and high-risk groups for EOC. The area under the curve (AUC), sensitivity and specificity were calculated for HE4, CA125 and ROMA for the diagnosis of ovarian cancer using receiver operating characteristic (ROC) analysis. RESULTS: The median CA125 and HE4 serum concentrations were significantly higher among EOC patients than in healthy females (both p<0.05) and those with a benign mass (both p<0.05). The pre-menopausal group included 36 benign cases (29 of which were classified by ROMA as low-risk with a specificity of 80.6%; 95% CI: 64.0%-91.8%), and 15 EOC (eight of which were classified by ROMA as high-risk, with a sensitivity of 53.3%; 95% CI: 26.6%-78.7%). The post-menopausal group enclosed 13 benign cases (11 of which were classified by ROMA as low-risk with a specificity of 84.6%; 95% CI: 54.6%-98.0%), and 40 EOC (33 of which were classified by ROMA as high-risk with a sensitivity of 82.5%; 95% CI: 67.2%-92.7%). In the pre-menopausal group, the AUC was 0.64 (p=0.12, 95% CI: 0.44-0.83) for CA125, 0.77 (p=0.003, 95% CI: 0.62-0.92) for HE4 and 0.77 (p=0.002, 95% CI: 0.63-0.92) for ROMA. In the post-menopausal group, the AUC was 0.84 (p=0.0003, 95% CI: 0.73-0.94) for CA125, 0.94 (p<0.0001, 95% CI: 0.88-0.99) for HE4 and 0.92 (p<0.0001, 95% CI: 0.85-0.99) for ROMA. CONCLUSIONS: The ROMA is a simple scoring system which shows excellent diagnostic performance for the detection of EOC in post-menopausal women, but not in pre-menopausal women. Moreover, the dual marker combination of HE4 and CA125 (ROMA) does not show better performance than HE4 alone.

Clinical associations of anti-CENP-B and anti-Scl70 antibody levels measured by multiplexed fluorescent microsphere immunoassay in systemic sclerosis.

The aim of this study was to assess the association between anti-CENP-B and anti-Scl70 antibody levels, measured by multiplexed fluorescent microsphere immunoassay, and the clinical features in patients affected by systemic sclerosis. Clinical evaluation of 80 scleroderma patients was performed in order to evaluate disease activity and organ involvement. Scleroderma-specific autoantibodies were detected using multiplexed fluorescent microsphere immunoassay. Unexpectedly, 11 patients resulted positive for both anti-Scl70 and anti-CENP-B antibodies; six cases showed a weak positivity for one of the two autoantibodies and a stronger positivity for the other one; five cases showed an intense positivity for both autoantibodies. This latter subgroup was excluded from the analysis of the associations between autoantibody levels and the clinical features. In the anti-CENP-B positive patients higher antibody levels were associated with a less extensive skin involvement in comparison with the cases affected by a more extensive skin involvement (521 +/- 208 vs 395 +/- 166 U/ml, respectively, P 0.038). In the anti-Scl70 positive patients autoantibody levels were directly correlated with skin involvement (P 0.018), showing higher levels in patients with a more extensive skin involvement in comparison with cases characterized by less extensive skin involvement (734 +/- 135 vs 490 +/- 183 U/ml, respectively, P 0.001). The findings of our study supports the association between autoantibody profile and disease severity in systemic sclerosis. In particular high levels of anti-Scl70 antibodies are associated with a worse cutaneous involvement, while high levels of anti-CENP-B antibodies seem to have a protective effect on skin manifestations.

The utility of serum human epididymis protein 4 (HE4) in patients with a pelvic mass.

AIM: Although CA125 is the most widely used cancer marker in the diagnostic approach of pelvic masses in women, its clinical usefulness is limited because it lacks expression of the antigen in the early stages of disease. The human epididymis protein 4 (HE4) is frequently over-expressed in ovarian cancer, whereas its expression in normal tissues, including the ovary, is low. The aim of this study was to assess the concentration of both HE4 and CA125 in patients with different forms of benign and malign pelvic masses. METHODS: The study population included 99 patients with gynecological cancer (46 ovarian, 39 endometrial, 14 cervical) and 40 affected by benign disease (22 endometriosis and 18 benign ovarian mass). Twelve control subjects were also included in the study. In all the patients, serum samples were collected on the day before scheduled surgery. RESULTS: The median CA125 and HE4 serum levels were significantly higher among ovarian cancer patients as compared with healthy subjects and with those with benign mass, cervical, and endometrial tumors. The receiver operating characteristics curve analysis on healthy controls and patients with ovarian cancers revealed that HE4 had a significantly higher area under the curve when compared with CA125 (0.99 vs. 0.91), with a sensibility and specificity of 98 and 100%, respectively. CONCLUSIONS: HE4 seems to be a promising ovarian cancer marker, and its measurement might improve the diagnostic approach to patients with pelvic masses.

Analytical evaluation of the novel Lumipulse G BRAHMS procalcitonin immunoassay.

OBJECTIVES: This study was designed to evaluate the analytical performance of the novel Lumipulse G1200 BRAHMS procalcitonin (PCT) immunoassay. DESIGN AND METHODS: This analytical evaluation encompassed the calculation of the limit of blank (LOB), limit of detection (LOD), functional sensitivity, intra- and inter-assay imprecision, confirmation of linearity and a comparison with the Vidas BRAHMS PCT assay. RESULTS: The LOB, LOD and functional sensitivity were 0.0010 ng/mL, 0.0016 ng/mL and 0.008 ng/mL, respectively. The total analytical imprecision was found to be 2.1% and the linearity was excellent (r=1.00) in the range of concentrations between 0.006-75.5 ng/mL. The correlation coefficient with Vidas BRAHMS PCT was 0.995 and the equation of the Passing and Bablok regression analysis was [Lumipulse G BRAHMS PCT]=0.76×[Vidas BRAHMS PCT]+0.04. The mean overall bias of Lumipulse G BRAHMS PCT versus Vidas BRAHMS PCT was -3.03 ng/mL (95% confidence interval [CI]: -4.32 to -1.74 ng/mL), whereas the mean bias in samples with PCT concentration between 0-10 ng/mL was -0.49 ng/mL (95% CI: -0.77 to -0.24 ng/mL). The diagnostic agreement was 100% at 0.5 ng/mL, 97% at 2.0 ng/mL and 95% at 10 ng/mL, respectively. CONCLUSIONS: These results attest that Lumipulse G BRAHMS PCT exhibits excellent analytical performance, among the best of the methods currently available on the diagnostic market. However, the significant bias compared to the Vidas BRAHMS PCT suggests that the methods cannot be used interchangeably.

A novel serum marker for biliary tract cancer: diagnostic and prognostic values of quantitative evaluation of serum mucin 5AC (MUC5AC).

BACKGROUND AND AIMS: Mucin 5AC (MUC5AC) is a glycoprotein found in different epithelial cancers, including biliary tract cancer (BTC). The aims of this study were to investigate the role of MUC5AC as serum marker for BTC and its prognostic value after operation with curative intent. PATIENTS AND METHOD: From January 2007 to July 2012, a quantitative assessment of serum MUC5AC was performed with enzyme-linked immunoassay in a total of 88 subjects. Clinical and biochemical data (including CEA and Ca 19-9) of 49 patients with BTC were compared with a control population that included 23 patients with benign biliary disease (BBD) and 16 healthy control subjects (HCS). RESULTS: Serum MUC5AC was greater in BTC patients (mean 17.93 ± 10.39 ng/mL) compared with BBD (mean 5.95 ± 5.39 ng/mL; P < .01) and HCS (mean 2.74 ± 1.35 ng/mL) (P < .01). Multivariate analysis showed that MUC5AC was related with the presence of BTC compared with Ca 19-9 and CEA: P < .01, P = .080, and P = .463, respectively. In the BTC group, serum MUC5AC ≥ 14 ng/mL was associated with lymph-node metastasis (P = .050) and American Joint Committee on Cancer and International Union for Cancer Control stage IVb disease (P = .047). Moreover, in patients who underwent operation with curative intent, serum MUC5AC ≥ 14 ng/mL was related to a worse prognosis compared with patients with lesser levels, with 3-year survival rates of 21.5% and 59.3%, respectively (P = .039). CONCLUSION: MUC5AC could be proposed as new serum marker for BTC. Moreover, the quantitative assessment of serum MUC5AC could be related to tumor stage and long-term survival in patients with BTC undergoing operation with curative intent.

Assessment of bile and serum mucin5AC in cholangiocarcinoma: diagnostic performance and biologic significance.

BACKGROUND: Recent studies have showed the efficacy of mucin5AC (MUC5AC) as a diagnostic and prognostic serum biomarker in biliary tract tumors. The aim of the present investigation was to improve the current knowledge on the biologic relevance of MUC5AC in malignant and benign biliary disorders by comparing its diagnostic performance in both bile and serum samples of patients with cholangiocarcinoma (CCA) or benign biliary disorders. METHODS: A quantitative determination of MUC5AC by enzyme-linked immunosorbent assay was performed in bile and serum specimens from 26 patients with extrahepatic CCA and 20 subjects with benign biliary disorders (10 with biliary stones and 10 with cholangitis). Verification analysis was made by immunoblot. RESULTS: MUC5AC of serum and biliary origin contributed to different extent to total levels of MUC5AC in the different groups of patients. In particular, the transition toward a greater degree of injury of bile duct epithelium was accompanied by a greater amount of MUC5AC in serum than in bile. The diagnostic performance of MUC5AC expressed as serum/bile ratio showed excellent diagnostic performance for differentiating CCA from cholangitis (area under the curve [AUC], 0.94; 95% CI, 0.86-1.00; P < .0001), CCA from biliary stones (AUC, 0.99; 95% CI, 0.98-1.00; P < .0001), as well as cholangitis from biliary stones (AUC, 0.93; 95% CI, 0.82-1.00; P = .001). CONCLUSION: These findings provide new insight into the biologic importance of MUC5AC in biliary disorders and suggest that combined assessment of MUC5AC in bile and serum with expression of data in terms of serum to bile ratio may improve the diagnostic performance of MUC5AC quantification in serum alone.

Hyaluronic acid: in vitro and in vivo analysis, biochemical properties and histological and morphological evaluation of injected filler.

BACKGROUND: No human model has emerged as an accepted standard to evaluate tissue filler longevity. OBJECTIVES: To validate a human model adequate to compare soft tissue filler degradation and tissue reaction. MATERIALS AND METHODS: We evaluated in 18 patients the persistence of hyaluronic acid (HA) filler injected into labial tissue analyzing hyaluronidase (HYAL) activity by means of in vitro and in vivo tests, MRI and histological and ultra-structural examination at 3 and 6 months postop. RESULTS: MRI examination revealed the presence of HA filler in a clear hyperintense area. Histology demonstrated fibroblast activation. The amount and the degradation rate of HYAL and HA did not show a linear correlation. CONCLUSION: MRI demonstrated the presence of HA in lip tissue even after 6 months. Biopsies at 3 months revealed tissue maturation and at 6 months confirmed the ability of HA to reorganize and integrate the extracellular matrix. The absence of linear correlation between HYAL and HA revealed that the result clinically is probably dependent on systemic factors which can determine HYAL activity and therefore HA longevity.

The role of resistin in colorectal cancer.

BACKGROUND: To date the role of resistin in colorectal cancer (CRC) is far from being elucidated. The aim of this study was to investigate the association between serum resistin levels and CRC in relation to known risk/protective factors including anthropometric, metabolic, inflammatory parameters as well as lifestyle individual characteristics. METHODS: 40 CRC patients and 40 controls were enrolled. Body weight, height, waist circumference and blood pressure were recorded. Fasting plasma glucose, lipids, C-reactive protein (CRP) and resistin levels were measured. Metabolic Syndrome (MS) was defined according to the harmonized definition. RESULTS: Resistin levels were significantly higher in CRC patients than in controls (p=0.028) and gradually increased with tumor stage progression (p=0.042). A high resistin level was statistically significant determinant of CRC after adjusting for age, sex, body mass index and lifestyle parameters (p=0.029). Resistin showed a strong association with CRP levels (p ≤ 0.0001). In stepwise regression analysis CRP remained the only independent predictor of both resistin levels (p=0.001) and CRC risk (p=0.021). CONCLUSIONS: These results clarify the nature of the association between resistin and CRC risk suggesting that the proinflammatory state of cancer, rather than the clinical diagnosis of CRC itself or its link with obesity and MS, may govern this association.

Very low levels of vitamin D in systemic sclerosis patients.

Vitamin D displays many extraosseous immunomodulatory effects. The aim of the study was to evaluate the level of vitamin D in patients with systemic sclerosis (SSc) and to analyze the associations between the concentration of the vitamin and clinical manifestations. In March-April 2009, 65 consecutive SSc patients underwent evaluation of vitamin D concentrations by the LIAISON immunoassay (normal 30-100 ng/ml). Serum levels between 10 and 30 ng/ml were classified as vitamin D insufficiency, while concentrations <10 ng/ml as vitamin D deficiency. None of the patients were receiving vitamin D supplementation at the time of or during the year prior to study entry. The mean level of vitamin D was 15.8 ± 9.1 ng/ml. Only three cases showed normal values; vitamin D insufficiency and deficiency were found in 43 and 19 cases, respectively. Patients with vitamin D deficiency showed longer disease duration (13.1 ± 6.8 versus 9.4 ± 5.5 years, P = 0.026), lower diffusing lung capacity for carbon monoxide (63.7 ± 12.4 versus 76.4 ± 20.2, P = 0.014), higher estimated pulmonary artery pressure (28.9 ± 9.9 versus 22.8 ± 10.4, P = 0.037) and higher values of ESR (40 ± 25 versus 23 ± 13 mm/h, P = 0.001) and of CRP (7 ± 7 and 4 ± 2 mg/l, P = 0.004) in comparison with patients with vitamin D insufficiency; moreover, late nailfold videocapillaroscopic pattern was more frequently found (52.6% versus 18.6%, P = 0.013). None of the patients showed evidence of overt mal-absorption. Low levels of vitamin D are very frequent in patients with SSc. Intestinal involvement is not likely the cause of vitamin D deficit; other factors such as skin hyperpigmentation and reduced sun exposition for psychological and social reasons may be implicated. Patients with vitamin D deficiency showed more severe disease in comparison with patients with vitamin D insufficiency, above all concerning lung involvement. Further trials are awaited to determine whether vitamin D could represent a modifiable factor able to interfere with SSc evolution.

Determination of ANA specificity using multiplexed fluorescent microsphere immunoassay in patients with ANA positivity at high titres after infliximab treatment: preliminary results.

To evaluate ANA specificity using the fully automated multiplexed fluorescent microsphere immunoassay in patients affected either by rheumatoid arthritis or ankylosing spondylitis who developed strong positivity for ANA as assessed by indirect immunofluorescent method on HEp-2 cells during infliximab treatment. Three men affected by ankylosing spondylitis and 12 women affected by rheumatoid arthritis who developed ANA positivity at high titres during infliximab treatment underwent the identification of ANA specificity by multiplexed fluorescent microsphere immunoassay; moreover anti-DNA and anti-ENA antibodies were tested by indirect immunofluorescence and ELISA method, respectively. In 4 out of 15 cases, the determination of ANA reactivity by multiplexed fluorescent microsphere immunoassay was also performed on the serum collected before infliximab administration. One patient affected by rheumatoid arthritis showed multiple ANA reactivities against SS-A, SS-B, RNP, Sm, Jo-1 and histones; one patient affected by ankylosing spondylitis resulted positive for the same autoantibodies, except for anti-Sm antibody. Moreover, two patients, one with rheumatoid arthritis and one with ankylosing spondylitis, showed single antibody specificity to SS-B and RNP, respectively. The remaining 11 cases did not show any positivity. Instead, all the patients resulted negative for anti-ENA antibodies by the ELISA method. In the four cases tested for ANA specificity by multiplexed fluorescent microsphere immunoassay before and after infliximab administration no difference was found. The search for anti-DNA antibody always resulted negative by both the traditional immunofluorescent assay and the novel technique. The use of multiplexed fluorescent microsphere immunoassay in patients treated with infliximab with ANA positivity at high titres allowed to find some ANA specificities which were not revealed by ELISA method. Nevertheless, the majority of patients resulted negative in spite of ANA positivity at high titres; the molecular target of ANA which develop after infliximab administration still remains to be identified.