Increased soluble (pro)renin receptor protein by autophagy inhibition in cultured cancer cells

(Pro)renin receptor ((P)RR) regulates the renin–angiotensin system and functions as an essential accessory subunit of vacuolar H⁺‐ATPase. There is accumulating evidence that shows close relationship between (P)RR and autophagy. Soluble (P)RR consisting of the extracellular domain of (P)RR is generated from (P)RR by proteolytic enzymes. The aim of the present study was to clarify the influence of autophagy inhibition on soluble (P)RR expression in cancer cells. Autophagy was inhibited by treatment of bafilomycin A1 or chloroquine in MCF‐7 and A549 cells for 72 hr. Western blot analysis showed that protein levels of soluble (P)RR were markedly elevated by autophagy inhibition, whereas no noticeable increases were observed in full‐length (P)RR. Secretion of soluble (P)RR into the medium was increased dose‐dependently by bafilomycin A1 or chloroquine. Autophagy inhibition was confirmed by enhanced accumulation of autophagy‐related proteins, LC3, p62 and LAMP1 in intracellular vesicles. Increased amount of soluble (P)RR by autophagy inhibition was decreased by site‐1 protease inhibitor, whereas no noticeable increase in site‐1 protease immunoreactivity was observed in cells with autophagy inhibition by immunocytochemistry. These findings suggest that soluble (P)RR protein accumulates by autophagy inhibition, possibly because of the reduced degradation of soluble (P)RR in the intracellular vesicles during autophagy inhibition.     

Detection of Norwalk Virus and Other Genogroup 1 Human Caliciviruses by a Monoclonal Antibody, RecombinantAntigen-Based Immunoglobulin M Capture Enzyme Immunoassay

Sera obtained from two groups of adult volunteers infected with Norwalk virus (NV) and two groups of patients involved in two natural outbreaks were tested for NV-reactive immunoglobulin M (IgM) by use of a monoclonal antibody, recombinant-antigen-based IgM capture enzyme immunoassay (EIA). No NV-reactive IgM was detected in the preinoculation sera of 15 volunteers, and 14 of 15 showed NV-reactive antibodies postinfection with NV. All of the volunteers showed IgG seroconversion to NV. In the outbreak studies, all 9 persons in one outbreak and 19 of 24 in another outbreak had NV-reactive IgM. In the first outbreak, only three of nine seroconverted to NV, which was likely due to late collection of acute-phase sera. In the second outbreak, 21 of 24 showed IgG seroconversion to NV. Sequencing of viruses isolated from five stool samples selected from those in the second outbreak showed that they were human calicivirus (HuCV) genogroup 1 viruses related, but not identical, to NV. In the volunteer studies, NV-reactive IgM was first detected 8 days postinoculation. The time of development of NV-reactive IgM antibodies in natural outbreaks was estimated to be similar to that found in the volunteer studies. Sera from three Hawaii virus-infected volunteers, four Snow Mountain virus patients, and 80 healthy individuals were negative for NV-reactive IgM, indicating test specificity for HuCV genogroup I infections. This capture IgM EIA is suitable for diagnosis of NV and other HuCV genogroup I infections and is especially useful when sera and fecal samples have not been collected early in the course of an outbreak.     

Functional activity of anti-Neisserial surface protein A monoclonal antibodies against strains of Neisseria meningitidis serogroup B

Neisserial surface protein A (NspA) is currently being investigated with humans as a candidate vaccine for the prevention of meningococcal disease. Although NspA is highly conserved, the ability of anti-NspA antibodies to bind to or elicit complement-mediated bactericidal activity against diverse Neisseria meningitidis serogroup B strains is controversial. To evaluate strain differences in NspA surface accessibility and susceptibility to bactericidal activity, we prepared murine immunoglobulin G2a anti-NspA monoclonal antibodies (MAbs) and evaluated their functional activity against 10 genetically diverse N. meningitidis serogroup B strains. By colony Western blot, all 10 strains expressed NspA as detected by one or more MAbs. By flow cytometry, two MAbs were found to bind to the bacterial surface of 6 of the 10 strains. In addition, two strains showed variable NspA surface accessibility for the MAbs despite being uniformly positive for NspA expression by colony Western blotting. Only 4 of the 10 strains were susceptible to anti-NspA complement-mediated bacteriolysis. Passively administered MAb protected infant rats from developing bacteremia after challenge with N. meningitidis serogroup B strain 8047 (surface binding positive, susceptible to anti-NspA bacteriolysis), was poorly protective against strain BZ232 (surface binding variable, resistant to bacteriolysis), and did not protect against strain M986 (surface binding negative, resistant to bacteriolysis). Finally, NspA does not appear to be critical for causing bacteremia, as an NspA knockout from strain 8047 was highly virulent in infant rats. Taken together, these findings suggest that an NspA-based vaccine will need to incorporate additional antigens to elicit broad protection against N. meningitidis serogroup B.     

Diagnosis of norwalk virus infection by indirect enzyme immunoassay detection of salivary antibodies to recombinant norwalk virus antigen

Simple diagnostic tests are needed for the detection of norovirus (NoV) outbreaks. Salivary antibody assays provide an attractive alternative to collecting and testing serum or stool samples. Antibodies to Norwalk virus (NV) in oral fluid samples were compared with NV antibodies in serum collected from 38 volunteers challenged with NV inoculum. Pre- and postchallenge (day 4, 8, 14, and 21) saliva and serum samples were examined by enzyme immunoassay (EIA) using recombinant NV antigen. Of 18 infected subjects (those who shed NV in stool or who demonstrated immunoglobulin G [IgG] seroconversion), 15 (83%) had > or =4-fold increases in NV-specific salivary IgA and 15 (83%) had > or =4-fold increases in NV-specific salivary IgG when prechallenge and postchallenge saliva samples were compared. When the results of the IgA and IgG assays were combined, all 18 infected subjects showed > or =4-fold increases in NV-specific salivary IgG or IgA postchallenge titers compared to their prechallenge titers. One of 19 uninfected subjects had a > or =4-fold increase in NV-specific salivary IgG. The sensitivity of the combined assay results was 100%, and the specificity was 95%. NV-specific salivary IgA titers peaked around 14 days postchallenge. NV-specific salivary IgG and serum IgG titers continued to rise through 21 days postchallenge. The application of this EIA to an elementary school outbreak indicated that 67% of the subjects with confirmed infections had >4-fold rises in anti-NoV IgA when an antigen in the same genetic cluster as the outbreak virus was used. This is the first documented mucosal antibody response to NoV in children. This EIA provides a useful approach for diagnosing NoV outbreaks.     

Longitudinal Development of Mother–Infant Interaction During the First Year of Life Among Mothers with Substance Abuse and Psychiatric Problems and Their Infants

The quality of mother–infant interaction during the first year may be hampered by maternal substance abuse and co-existing non-optimal factors such as psychiatric problems and difficult relational experiences. In the present study three groups of women were recruited during pregnancy: One group with substance abuse problems from residential treatment centers (n = 28), a second group from psychiatric outpatient treatment centers (n = 22), and a third group from well-baby clinics (n = 30). Four maternal optimality indexes were assessed (substance abuse, psychiatric problems, relational experiences and SES). Mother–infant interaction was observed at 3 and 12 months. The substance abuse group showed the most disturbed mother–infant interaction at 12 months. Low maternal optimality as well as impairments in maternal affective involvement at 3 months influenced negatively both on infant and dyadic affective behavior in interaction at 12 months. Long-term interventions are needed to promote affective reciprocity among mother–baby pairs with low optimality.     

Correction to: Pleiotropic effect of erythropoiesis‑stimulating agents on circulating endothelial progenitor cells in dialysis patients

Clinical and Experimental Nephrology -     

BMK1 is activated in glomeruli of diabetic rats and in mesangial cells by high glucose conditions

BACKGROUND: High glucose causes renal cell injury through various signal transduction pathways, including mitogen-activated protein (MAP) kinases cascades. Big MAP kinase 1 (BMK1), also known as extracellular signal-regulated kinase 5 (ERK5), is a recently identified MAP kinase family member and was reported to be sensitive to osmotic and oxidative stress. However, the role of BMK1 in diabetic nephropathy has not been elucidated yet. METHODS: We investigated whether BMK1 is activated in the glomeruli of Otsuka Long Evans Tokushima Fatty (OLETF) rats, a model of type 2 diabetes mellitus in comparison with the control Long Evans Tokushima Otsuka (LETO) rats. We also examined the effect of high glucose on BMK1 activity in cultured rat mesangial cells. RESULTS: BMK1 and ERK1/2 but not p38 were activated in the glomeruli of OLETF rats, which showed diabetic nephropathy at 52 weeks of age. High glucose, in addition to a high concentration of raffinose, caused rapid and significant activation of BMK1 in rat mesangial cells. MAP kinase/ERK kinase (MEK) inhibitors, U0126 and PD98059, both inhibited BMK1 activation by high glucose in a concentration-dependent manner. Protein kinase C (PKC) inhibition by GF109203X and PKC down-regulation with long-time phorbol myristate acetate (PMA) treatment both inhibited BMK1 and Src kinase activation. Src kinase inhibitors, herbimycin A and PP2, also inhibited high glucose-induced BMK1 activation. PKC inhibitors, Src inhibitors and MEK inhibitors, all inhibited cell proliferation by high glucose. Finally, transfection of dominant-negative MEK5, which is an upstream regulator of BMK1, abolished the BMK1-mediated rat mesangial cell proliferation stimulated by high glucose. CONCLUSION: In the present study, we demonstrated that high glucose activates BMK1 both in vivo and in vitro. It was suggested that high glucose induces PKC- and c-Src-dependent BMK1 activation. It could not be denied that BMK1 activation is induced through an osmotic stress-sensitive mechanism. BMK1-mediated mesangial cell growth may be involved in the pathogenesis of diabetic nephropathy.     

Transrectal ultrasound biopsy of the prostate: does it still have a role in prostate cancer diagnosis?

Transrectal ultrasound (TRUS) guided biopsy of the prostate has been a standard diagnostic approach for prostate cancer over the past thirty years. Today, the role of TRUS biopsy is being challenged by transperineal (TP) prostate biopsy due to concerns over the safety and diagnostic yield of TRUS biopsy. TRUS biopsy still offers a convenient, reliable and accessible tool for diagnosing prostate cancer in the majority of patients. It continues to play a role in prostate cancer diagnosis, especially where hospital resource allocation is limited, including the public sector. TRUS biopsy has low rates of severe complications, although there remains room for improvement in current practice to improve the tolerability and reduce the incidence of post-biopsy infection.