John J. Chang

Renalase Prevents AKI Independent of Amine Oxidase Activity

AKI is characterized by increased catecholamine levels and hypertension. Renalase, a secretory flavoprotein that oxidizes catecholamines, attenuates ischemic injury and the associated increase in catecholamine levels in mice. However, whether the amine oxidase activity of renalase is involved in preventing ischemic injury is debated. In this study, recombinant renalase protected human proximal tubular (HK-2) cells against cisplatin- and hydrogen peroxide-induced necrosis. Similarly, genetic depletion of renalase in mice (renalase knockout) exacerbated kidney injury in animals subjected to cisplatin-induced AKI. Interestingly, compared with the intact renalase protein, a 20-amino acid peptide (RP-220), which is conserved in all known renalase isoforms, but lacks detectable oxidase activity, was equally effective at protecting HK-2 cells against toxic injury and preventing ischemic injury in wild-type mice. Furthermore, in vitro treatment with RP-220 or recombinant renalase rapidly activated Akt, extracellular signal-regulated kinase, and p38 mitogen-activated protein kinases and downregulated c-Jun N-terminal kinase. In summary, renalase promotes cell survival and protects against renal injury in mice through the activation of intracellular signaling cascades, independent of its ability to metabolize catecholamines, and we have identified the region of renalase required for these effects. Renalase and related peptides show potential as therapeutic agents for the prevention and treatment of AKI.

Inhibition of Na(+)-K(+)-2Cl(-) cotransport by mercury.

Mercury alters the function of proteins by reacting with cysteinyl sulfhydryl (SH(-)) groups. The inorganic form (Hg(2+)) is toxic to epithelial tissues and interacts with various transport proteins including the Na(+) pump and Cl(-) channels. In this study, we determined whether the Na(+)-K(+)-Cl(-) cotransporter type 1 (NKCC1), a major ion pathway in secretory tissues, is also affected by mercurial substrates. To characterize the interaction, we measured the effect of Hg(2+) on ion transport by the secretory shark and human cotransporters expressed in HEK-293 cells. Our studies show that Hg(2+) inhibits Na(+)-K(+)-Cl(-) cotransport, with inhibitor constant (K(i)) values of 25 microM for the shark carrier (sNKCC1) and 43 microM for the human carrier. In further studies, we took advantage of species differences in Hg(2+) affinity to identify residues involved in the interaction. An analysis of human-shark chimeras and of an sNKCC1 mutant (Cys-697-->Leu) reveals that transmembrane domain 11 plays an essential role in Hg(2+) binding. We also show that modification of additional SH(-) groups by thiol-reacting compounds brings about inhibition and that the binding sites are not exposed on the extracellular face of the membrane.

Identification of a receptor for extracellular renalase.

Background An increased risk for developing essential hypertension, stroke and diabetes is associated with single nucleotide gene polymorphisms in renalase, a newly described secreted flavoprotein with oxidoreductase activity. Gene deletion causes hypertension, and aggravates acute ischemic kidney (AKI) and cardiac injury. Independent of its intrinsic enzymatic activities, extracellular renalase activates MAPK signaling and prevents acute kidney injury (AKI) in wild type (WT) mice. Therefore, we sought to identity the receptor for extracellular renalase. Methods and Results RP-220 is a previously identified, 20 amino acids long renalase peptide that is devoid of any intrinsic enzymatic activity, but it is equally effective as full-length recombinant renalase at protecting against toxic and ischemic injury. Using biotin transfer studies with RP-220 in the human proximal tubular cell line HK-2 and protein identification by mass spectrometry, we identified PMCA4b as a renalase binding protein. This previously characterized plasma membrane ATPase is involved in cell signaling and cardiac hypertrophy. Co-immunoprecipitation and co-immunolocalization confirmed protein-protein interaction between endogenous renalase and PMCA4b. Down-regulation of endogenous PMCA4b expression by siRNA transfection, or inhibition of its enzymatic activity by the specific peptide inhibitor caloxin1b each abrogated RP-220 dependent MAPK signaling and cytoprotection. In control studies, these maneuvers had no effect on epidermal growth factor mediated signaling, confirming specificity of the interaction between PMCA4b and renalase. Conclusions PMCA4b functions as a renalase receptor, and a key mediator of renalase dependent MAPK signaling.

Renalase expression by melanoma and tumor associated-macrophages promotes tumor growth through a STAT3-mediated mechanism

To sustain their proliferation, cancer cells overcome negative-acting signals that restrain their growth and promote senescence and cell death. Renalase (RNLS) is a secreted flavoprotein that functions as a survival factor after ischemic and toxic injury, signaling through the plasma calcium channel PMCA4b to activate the PI3K/AKT and MAPK pathways. We show that RNLS expression is increased markedly in primary melanomas and CD163(+) tumor-associated macrophages (TAM). In clinical specimens, RNLS expression in the tumor correlated inversely with disease-specific survival, suggesting a pathogenic role for RNLS. Attenuation of RNLS by RNAi, blocking antibodies, or an RNLS-derived inhibitory peptide decreased melanoma cell survival, and anti-RNLS therapy blocked tumor growth in vivo in murine xenograft assays. Mechanistic investigations showed that increased apoptosis in tumor cells was temporally related to p38 MAPK-mediated Bax activation and that increased cell growth arrest was associated with elevated expression of the cell-cycle inhibitor p21. Overall, our results established a role for the secreted flavoprotein RNLS in promoting melanoma cell growth and CD163(+) TAM in the tumor microenvironment, with potential therapeutic implications for the management of melanoma. Cancer Res; 76(13); 3884-94. ©2016 AACR.

Risk of deep vein thrombosis in patients with cellulitis and erysipelas: a systematic review and meta-analysis.

INTRODUCTION The occurrence of deep vein thrombosis (DVT) is often considered in patients with cellulitis and erysipelas because of the common presentation of unilateral limb swelling, erythema and pain. Different authors however have reached different conclusions about the prevalence of DVT in these patients and for the need for compression ultrasound (CUS). The purpose of this study is to determine the prevalence of DVT in patients with cellulitis and erysipelas, and inform the utility of CUS. METHODS A systematic literature search was conducted of Medline and Cochrane for studies that reported groups of patients with cellulitis or erysipelas who had CUS to evaluate for DVT. Study quality assessment was based on the Newcastle-Ottawa Quality Assessment Scale for Cohort Studies. The incidence rates from the included studies were pooled using a random-effects model to calculate an overall DVT rate. Individual and pooled DVT rates with corresponding upper and lower limits were graphed as a forest plot. Between-study heterogeneity was estimated using the I(2) statistic. RESULTS Nine studies were included totaling 1054 patients with cellulitis or erysipelas with 18 DVTs. The overall pooled incidence rate was 2.1% (95% confidence interval, 0.5%-9.1%) for proximal DVT and 3.1% (95% confidence interval, 1.9%-4.9%) for any DVT. When analyzed separately, the pooled incidence rate for the three retrospective studies was 1.1% (95% CI, 0.6%-2.2%), while the rate for the six prospective studies was 7.8% (95% CI, 4.2%-14.2%). CONCLUSION The risk of DVT in cellulitis and erysipelas is low compared to the average risk of patients referred for CUS and comparable to low risk patients as determined by the commonly employed Wells criteria.

Extracellular renalase protects cells and organs by outside-in signalling

Renalase was discovered as a protein synthesized by the kidney and secreted in blood where it circulates at a concentration of approximately 3–5 μg/ml. Initial reports suggested that it functioned as an NAD(P)H oxidase and could oxidize catecholamines. Administration of renalase lowers blood pressure and heart rate and also protects cells and organs against ischaemic and toxic injury. Although renalases protective effect was initially ascribed to its oxidase properties, a paradigm shift in our understanding of the cellular actions of renalase is underway. We now understand that, independent of its enzymatic properties, renalase functions as a cytokine that provides protection to cells, tissues and organs by interacting with its receptor to activate protein kinase B, JAK/STAT, and the mitogen‐activated protein kinase pathways. In addition, recent studies suggest that dysregulated renalase signalling may promote survival of several tumour cells due to its capacity to augment expression of growth‐related genes. In this review, we focus on the cytoprotective actions of renalase and its capacity to sustain cancer cell growth and also the translational opportunities these findings represent for the development of novel therapeutic strategies for organ injury and cancer.

Diagnostic accuracy of methicillin-resistant Staphylococcus aureus nasal colonization to predict methicillin-resistant S aureus soft tissue infections

Nasal methicillin-resistant Staphylococcus aureus (MRSA) testing at admission to the hospital was found to have a positive likelihood ratio of 8.5 and a negative likelihood ratio of 0.41 for predicting MRSA soft tissue infections. The clinical utility of this test depends on the prevalence of MRSA infection. In high prevalence populations, nasal MRSA is useful to rule in MRSA infections. In low prevalence populations it may be useful to rule out infections.

Overuse of compression ultrasound for patients with lower extremity cellulitis.

BACKGROUND Compression ultrasound (CUS) is often ordered in hospitalized patients with cellulitis to assess for deep vein thrombosis (DVT). Despite this common practice, the rate of use and utility of CUS has not been well described. METHODS We conducted a retrospective cohort study of adult patients with lower extremity cellulitis hospitalized between October 1, 2008 and September 30, 2013 at an academic medical center. Cases meeting inclusion criteria were reviewed for the use of CUS, the indication for CUS, the occurrence of DVT, and the 3 month follow-up occurrence of DVT after discharge. RESULTS A total of 239 patients were identified using ICD-9 coding data with a discharge diagnosis of cellulitis or abscess of leg. Of these, 183 met criteria for inclusion in the study, 133 of whom had CUS to assess for DVT (73%). Of the 133 who received CUS, 11 studies found DVTs (8%). Of the 11 DVTs, 8 had been previously diagnosed, and 3 were new. Of the new DVTs, only one was ipsilateral to the leg with cellulitis. CONCLUSION Most patients admitted with lower extremity cellulitis received CUS to assess for DVT. Despite this common practice, the rate of acute ipsilateral DVT was low and matched the rate of acute contralateral DVT. Previously diagnosed DVTs were commonly re-imaged. Overall the use of CUS had minimal impact on patient management and the routine use of CUS to assess for DVT in hospitalized patients with cellulitis appears to be unnecessary.

Single-chip Integration of SRAM and Non-volatile Memory using Bit-line Sharing

A new memory architecture integrating SRAM and Flash within the same array is presented and demonstrated in a standard 0.25mum CMOS process with a memory access time of 20ns. Differential pair Flash cells with low programming current share the same bit-lines as SRAM cells within the same array. This enables row-to-row transfer of data between Flash and SRAM cells as well as access data through I/O directly, in return improving speed and lowering power. Area is saved through the shared usage of column decoding, sense-amplifier, and write-driver circuitry

Clinical characteristics and outcomes of veterans hospitalised with purulent soft tissue infections with and without systemic signs of infection

Abstract Objective To describe the frequency of systemic inflammatory response syndrome (SIRS) criteria in a cohort of patients hospitalised with purulent soft tissue infections and to determine their impact on clinical characteristics, microbiology and outcomes. Methods Retrospective cohort study of adults hospitalised at the West Haven Veteran’s Hospital with purulent soft tissue infections between 1 October 2008 and 30 September 2013. Results A total of 173 patients were included with purulent soft tissue infections; 60 patients had no SIRS, 48 had one SIRS and 65 had ≥ 2 SIRS. Most clinical characteristics were similar between the different SIRS groups, although patients with SIRS were more likely to have severe sepsis and acute kidney injury and to already be on antibiotics at the time of hospitalisation. The microbiology of the infections was similar between SIRS groups. All patients received parenteral antibiotics when admitted and the majority of patients in all SIRS categories received antibiotics with broad Gram-negative activity. Outcomes were generally benign for all SIRS groups, although patients with SIRS had a longer length of stay and a trend towards more bacteremia. Conclusions SIRS are common in patients hospitalised with purulent soft tissue infections, although one third had no systemic signs of infection. Severe sepsis and septic shock are rare. Clinical characteristics, microbiology and antibiotic use are similar among patients in different SIRS groups, although the group without SIRS had a shorter hospitalisation and no episodes of bacteremia. Over-use of antibiotics is common in all SIRS categories.