Circulating microRNAs in serum: novel biomarkers for patients with bladder cancer?

Purpose

Recent studies indicate that circulating microRNAs in serum/plasma are a novel class of non-invasive biomarkers with diagnostic and prognostic information. So far, circulating microRNAs have not been analyzed in patients with bladder cancer.

Methods

We collected serum from patients with non-muscle invasive bladder cancer (NMIBC), muscle invasive bladder cancer (MIBC) and non-malignant urological disease. Total RNA was isolated from 400 μl of serum using the mirVana PARIS Kit; the artificial cel-miR-39 was spiked-in prior to RNA isolation to control different RNA isolation efficiencies. Quantitative real-time PCR was applied to measure the levels of 22 microRNAs upregulated in BCA tissue (miR-15a, miR-18a, miR-21, miR-93, miR-96, miR-103, miR-130b, miR-135b, miR-141, miR-182, miR-183, miR-190, miR-191, miR-200b, miR-422b, miR-425, miR-449b, miR-601, miR-639, miR-644, miR-649 and miR-1233) in the marker identification cohort (NMIBC, n = 11, MIBC, n = 10; controls, n = 10). The most promising serum microRNAs were tested in a validation cohort (NMIBC, n = 65, MIBC, n = 61; controls, n = 105).

Results

The RNA recovery was similar in patients with NMIBC, MIBC and control subjects. The analysis of serum microRNA levels in the marker identification cohort indicated that serum miR-141 and miR-639 levels were increased in bladder cancer patients compared to CTRL. The analysis of these miR-141 and miR-639 in the validation cohort demonstrated that microRNA levels were similar in bladder cancer patients and control subjects. Furthermore, microRNA levels were not correlated with clinicopathological parameters (pT-stage, metastasis, grading).

Conclusions

The analysis of serum miR-141 and miR-639 levels does not seem to be helpful in the diagnosis or prognosis of BCA.     

Urinary α and β C-Telopeptides of Collagen I: Clinical Implications in Bone Remodeling in Patients with Anorexia Nervosa

Fragments derived from degradation of type I collagen C-telopeptide (CTX) can be nonisomerized (α) or β-isomerized (β) depending on the age of bone; i.e., mainly the α form is derived from new bone and the β form from old bone. We have studied 41 female patients with anorexia nervosa (AN), aged 18.5 ± 2.2 years (range 16–24 years), and with an evolution time between 1.5 and 11 years, and 31 healthy control females (C), with a mean age of 19 ± 2.3 years (range 16–24 years). The AN patients showed a significant decrease in bone mass, with a mean Z-score of bone mineral density (BMD) of −3.2 ± 0.8 (range −0.9 to −5.4). The aim of our study was to determine the levels of urinary α- and β-CTX markers of bone resorption, the α/β ratio (α/β), and the level of bone alkaline phosphatase (bAP), a biochemical marker of bone formation, in order to relate them to the degree of osteopenia and the status of bone remodeling. Statistical analysis was by the Mann–Whitney test. The degree of osteopenia correlated with bAP levels (p= 0.0027) but not with the other parameters. Patients with AN were divided into three groups according to their levels of bAP: high (H), normal (N) or low (L). We found that BMD was significantly lower, and α- and β-CTX were significantly higher, in groups H and N than in group L. Bone AP correlated significantly with α-CTX (p= 0.0042) and α/β (0.0095) in the controls, but not with β-CTX, while in AN patients bAP correlated with β-CTX (p= 0.0000) and with α-CTX (p= 0.022) but not with the α/β ratio. The ratio CTX/bAP (resorption/formation) was similar in AN patients and controls. It is concluded that: (1) patients with AN have a high degree of osteopenia which correlated with bAP levels; (2) urinary CTX fragments found in AN patients seem to come mainly from old bone (β-CTX), while CTX found in healthy adolescent control females come from new bone (α-CTX). For this reason, α-CTX is more suitable than β-CTX for measuring bone resorption in controls and β-CTX is more suitable in patients with AN; (3) the resorption/formation ratio (CTX/bAP) was similar in AN patients and controls. From points (2) and (3) it is possible to suggest that, although bAP reflects bone formation in control females, this marker does not reflect effective bone mineralization in AN patients, a similar feature to that of patients with osteomalacia. Received: 20 January 1999 / Accepted: 28 May 1999     

Gender- and Compartment-Specific Bone Loss in C57BL/6J Mice: Correlation to Season?

Seasonal variation in bone mineral density (BMD) has been documented in humans, and has been attributed to changes in 25-hydroxyvitamin D [25(OH)D] synthesis. To test the hypothesis that seasonal changes in bone mass occur in laboratory mice, we measured body composition, femoral bone phenotypes, and serum bone markers in 16-wk-old male and female C57BL/6 (B6) mice during the summer (June–August) and winter (December–February) months at The Jackson Laboratory in Bar Harbor, Maine. Both male and female B6 mice had higher volumetric BMD in the summer than winter. Females showed reduced trabecular bone, whereas males showed changes in bone volume. Males, but not females, had higher insulin-like growth factor 1 in summer than in winter, and only males showed an increase in body weight during the winter. No seasonal differences in serum TRAP5b, osteocalcin, or 25(OH)D were noted for either sex. We conclude that seasonal variation in skeletal and body composition parameters in B6 mice is significant and must be considered when performing longitudinal phenotyping of the skeleton. Further studies are needed to determine the environmental factors that cue seasonal changes in body composition and the mechanisms that produce these changes.     

Matrix Delivery Transdermal 17β-Estradiol for the Prevention of Bone Loss in Postmenopausal Women

A total of 277 early postmenopausal women were enrolled in this placebo-controlled 2-year study to examine the efficacy of a matrix transdermal 17β-estradiol system, at three different dosages (25, 50 and 75 mg/day) combined with sequential oral dydrogesterone 20 mg/day, in preventing bone loss. At 2 years, the difference from placebo in percentage change from baseline of L1–4 lumbar spine bone mineral density (BMD) (assessed by dual-energy X-ray absorptiometry) was 4.7%± 0.7% with estradiol 25 mg/day, 7.3%± 0.7% with estradiol 50 mg/day and 8.7%± 0.7% with estradiol 75 mg/day (all values mean ± SEM). There were also significant increases in femoral neck, trochanter and total hip BMD with all doses of estradiol compared with placebo. Additionally, most patients had a significant gain (increase greater than 2.08%) in lumbar spine bone mass compared with placebo. Patients who received estradiol also experienced clinically significant and dose-related decreases in total serum osteocalcin, serum bone alkaline phosphatase and urinary C-telopeptide, with all three markers of bone turnover returning to premenopausal levels. Estradiol was well tolerated during the 2-year treatment period. Transdermal estradiol is effective and well tolerated at dosages between 25–75 mg/day in the prevention of bone loss in postmenopausal women; 25 mg/day offers an effective option for those women who cannot tolerate higher doses. Received: 30 June 1998 / Accepted: 22 September 1998     

Editorial Commentary: Is Arthroscopic Bone Grafting Nearly Equivalent to Open Bone Grafting for Glenoid Bone Defects in Recurrent Anterior Shoulder Instability?

Arthroscopic techniques are an emerging technology to deal with glenoid bone defects in patients with anterior shoulder instability, and improvements are being made to safely minimize the risk of injury to the anterior neurovascular structures including the axillary nerve. Arthroscopic glenoid reconstruction is a technically demanding procedure, but it does have promising short-term outcomes. I truly like the concept of anterior (and also posterior) bone grafting for defects of the glenoid, including the arthroscopic Latarjet. A free bone graft (iliac crest, distal tibia) is part of a logical surgical learning curve progression to treat bone defects from an arthroscopic standpoint. Before performing an arthroscopic Latarjet, I might suggest looking at performing free bone block fixation arthroscopically. But, for now, I still enjoy the success of an open bone grafting procedure and will continue to use open as my primary bone grafting (Latarjet, distal tibia, iliac crest), so as to optimize the position of the graft for successful long-term outcomes. We look forward to seeing more of the authors' work and a longer term follow-up of these patients to clearly delineate the development of osteoarthrosis, recurrent instability, and long-term stability of the bone graft and shoulder joint function.     

Pancreatic β-cell dysfunction in type 2 diabetes: Implications of inflammation and oxidative stress

Insulin resistance and pancreatic β-cell dysfunction are major pathological mechanisms implicated in the development and progression of type 2 diabetes(T2D). Beyond the detrimental effects of insulin resistance, inflammation and oxidative stress have emerged as critical features of T2D that define β-cell dysfunction. Predominant markers of inflammation such as C-reactive protein, tumor necrosis factor alpha, and interleukin-1β are consistently associated with β-cell failure in preclinical models...     

Editorial Commentary: Glenoid Bone Loss Measurements in Shoulder Instability—Precise but Not Accurate

Glenoid defects are important to consider when choosing the surgical stabilization technique in shoulder instability patients. Several measurement methods to determine the extent of glenoid bone loss have been proposed and their reliability or precision proved. However, it must be considered that these defect extent measurements are only surrogate parameters trying to express the loss of biomechanical stability generated by a glenoid defect, which in fact they do not do accurately. Current defect measurement techniques are either linear based (1-dimensional) or area based (2-dimensional) but do not take into account the 3-dimensional shape of the glenoid concavity, which creates stability by means of the concavity-compression effect. Furthermore, none of the current measurement methods take into account the native glenoid concavity shape, which significantly differs between patients and therefore also affects the biomechanical consequence a glenoid defect generates. To improve the accuracy of current glenoid defect measurement techniques in expressing the loss of biomechanical stability generated by a glenoid defect, measurements should take into account the concave shape of the glenoid (3-dimensional measurements) and account for the baseline shape of the native glenoid (4-dimensional measurements).     

Editorial Commentary: Unsolved Problems in Shoulder Instability—The Dilemma of Bone Loss!

Bone loss in the setting of shoulder instability is an unsolved problem. Procedures restoring bone stock on the glenoid side vary from minimally invasive arthroscopic techniques of arthroscopic bone block procedures to traditional transposition of the coracoid or bone transplant. These techniques are evolving, and several tips and tricks have been optimized to improve outcomes and to reduce the risk of complications, even though the most terrible complication of transposition of the coracoid remains osteoarthritis in the young adult. The major innovation in the past few years has been the use of remplissage. The aim of our surgical procedures is to restore the anatomy as much as possible. Therefore, gentle handling of bone defects on both the humeral and glenoid sides by means of an arthroscopic bone block and combined remplissage seems to be the future of our surgical procedures.     

The intuitive case for β-blockers in patients with ESRD

Sudden cardiac death (SCD) is common in dialysis patients accounting for up to 25% of all-cause mortality. Unlike in the general population, occlusive coronary artery disease is implicated in a minority of these deaths. Activation of the sympathetic nervous system is prevalent in the dialysis population and may underlie this high rate of SCD. β-blockers reduce SCD in the general population and, given their mode of action, β-blockers would seem to be an ideal class of agents to prevent SCD in dialysis patients. In this review, we will explore the etiology of SCD in dialysis patients and discuss the evidence supporting the use of β-blockers in patients with ESRD. We will also examine potential impediments to the use β-blocker in the dialysis population and outline directions for future trials in this area.     

SH2B1 in β-Cells Regulates Glucose Metabolism by Promoting β-Cell Survival and Islet Expansion

IGF-1 and insulin promote β-cell expansion by inhibiting β-cell death and stimulating β-cell proliferation, and the phosphatidylinositol (PI) 3-kinase/Akt pathway mediates insulin and IGF-1 action. Impaired β-cell expansion is a risk factor for type 2 diabetes. Here, we identified SH2B1, which is highly expressed in β-cells, as a novel regulator of β-cell expansion. Silencing of SH2B1 in INS-1 832/13 β-cells attenuated insulin- and IGF-1–stimulated activation of the PI 3-kinase/Akt pathway and increased streptozotocin (STZ)-induced apoptosis; conversely, overexpression of SH2B1 had the opposite effects. Activation of the PI 3-kinase/Akt pathway in β-cells was impaired in pancreas-specific SH2B1 knockout (PKO) mice fed a high-fat diet (HFD). HFD-fed PKO mice also had increased β-cell apoptosis, decreased β-cell proliferation, decreased β-cell mass, decreased pancreatic insulin content, impaired insulin secretion, and exacerbated glucose intolerance. Furthermore, PKO mice were more susceptible to STZ-induced β-cell destruction, insulin deficiency, and hyperglycemia. These data indicate that SH2B1 in β-cells is an important prosurvival and proproliferative protein and promotes compensatory β-cell expansion in the insulin-resistant state and in response to β-cell stress.     

Bone marrow fibrosis with fibrocytic and immunoregulatory responses induced by β-catenin activation in osteoprogenitors

Wnt/β-catenin signaling has been reported to contribute to the development of bone fibrous dysplasia. However, it remains unclear whether fibrocytes and immune cells are involved in this β-catenin-mediated bone marrow fibrosis. In this study, we showed that constitutive activation of β-catenin by Col1a1-Cre (3.6-kb) exhibited bone marrow fibrosis, featured with expanded populations of fibrocytes, myofibroblasts and osteoprogenitors. Lineage tracing and IHC examinations showed that Col3.6-Cre display Cre recombinase activity not only in osteoprogenitors, but also in monocyte-derived fibrocytes in the endosteal niches of bones. Additionally, β-catenin stimulated the secretion of cytokines and pro-fibrotic signals in bone marrow, including GM-CSF, TGFβ1 and VEGF. Consequently, the frequency of differentiated immature monocyte-derived dendritic cells and naïve T cells was markedly increased in the mutant bone marrow. These phenotypes were quite different from those following β-catenin activation in mature osteoblasts driven by Col1a1-Cre (2.3-kb). Our findings suggested that a conserved pro-fibrotic signal cascade might underlie β-catenin-mediated bone marrow fibrosis, involving TGFβ1-enhanced fibrocyte activation and immunoregulatory responses. This study might shed new light on the understanding and development of a therapeutic strategy for bone fibrous dysplasia.     

Treatment with β2-agonist for severe bladder dysfunction due to high-dose irradiation after radical hysterectomy

A case of severe bladder dysfunction due to a high-dose irradiation of 180 Gy after radical hysterectomy for cervical cancer is presented. The patient had suffered from urinary disturbances for 11 years after operation and radiation therapy. The results of urodynamic studies were low capacity, low compliance and low urethral pressure. The ₂-agonist mabuterol was administered for 3 years. Bladder capacity, compliance and urethral closure pressure improved significantly after the treatment. The voided volume increased significantly to twice the pretreatment state. Frequency, incontinence and urine loss by the 60-minute padweighing test decreased significantly to one-third to one-fifth of the pretreatment state. The patient is doing well without any side effects 3 years after initiation of the treatment.     

Bioglass ®45S5 Stimulates Osteoblast Turnover and Enhances Bone Formation In Vitro: Implications and Applications for Bone Tissue Engineering

We investigated the concept of using bioactive substrates as templates for in vitro synthesis of bone tissue for transplantation by assessing the osteogenic potential of a melt-derived bioactive glass ceramic (Bioglass 45S5) in vitro. Bioactive glass ceramic and bioinert (plastic) substrates were seeded with human primary osteoblasts and evaluated after 2, 6, and 12 days. Flow cytometric analysis of the cell cycle suggested that the bioactive glass-ceramic substrate induced osteoblast proliferation, as indicated by increased cell populations in both S (DNA synthesis) and G2/M (mitosis) phases of the cell cycle. Biochemical analysis of the osteoblast differentiation markers alkaline phosphatase (ALP) and osteocalcin indicated that the bioactive glass-ceramic substrate augmented osteoblast commitment and selection of a mature osteoblastic phenotype. Scanning electron microscopic observations of discrete bone nodules over the surface of the bioactive material, from day 6 onward, further supported this notion. A combination of fluorescence, confocal, transmission electron microscopy, and X-ray microprobe (SEM-EDAX) examinations revealed that the nodules were made of cell aggregates which produced mineralized collagenous matrix. Control substrates did not exhibit mineralized nodule formation at any point studied up to 12 days. In conclusion, this study shows that Bioglass 45S5 has the ability to stimulate the growth and osteogenic differentiation of human primary osteoblasts. These findings have potential applications for tissue engineering where this bioactive glass substrate could be used as a template for the formation of bioengineered bone tissue.