A Novel Technique to Minimize Contamination for Cervical Cancer Surgery Patients.

The Laparoscopic Approach to Cervical Cancer (LACC) trial changed the surgical management of cervical cancer worldwide. It was a multinational phase III clinical trial that reported lower survival and higher rate of abdominopelvic recurrences in minimally invasive surgery (MIS) than those of open surgery after hysterectomy. It is possible that tumor exposure to the peritoneal cavity in the MIS arm may account for these differences. We propose a novel technique to minimize peritoneal contamination of malignant cells present at the cervical os by placing a vaginal cerclage abdominally to create a seal at the apex of the vagina during MIS radical hysterectomy. The 2 patients in this work remain healthy and disease-free more than 18 months after surgery using this novel technique. We intend this work to serve as a platform both for offering a safe alternative to the open approach supported by the LACC trial and, most importantly, for promoting discussion of the results of the LACC trial and further research on surgical techniques in the treatment of cervical cancer. MIS has repeatedly been shown to have lower rates of infection, fewer complications, and shorter hospital stays while providing oncologic care that is noninferior to open approach.

Current Trends in Unilateral Cleft Lip Care: A 10-Year Update on Practice Patterns.

BACKGROUND: The care of unilateral cleft lip (UCL) patients is extremely variable. Historical benchmarks for perioperative and intraoperative choices by cleft surgeons were produced by Sitzman et al (Plast Reconstr Surg. 2008;121:261e-270e) in 2005. However, emerging data and cleft lip repair methods around this period were not captured by this study. The aim of this study was to update the current practice patterns of cleft lip surgeons. METHODS: An electronic survey was distributed to surgeons in the American Cleft Palate Association. Demographic data about the surgeon were collected as well as their choices regarding perioperative and intraoperative cleft lip care. RESULTS: Eighty-six surgeons responded to the survey. Nearly 40% of surgeons have changed their technique for UCL repair with Fisher anatomical subunit repair gaining significant popularity. Nasoalveolar molding is also being used more frequently (41% vs 22%). At the time of the cleft lip repair, closure of the nasal floor is occurring in 83.1% of patients and primary cleft rhinoplasty is being performed routinely 57% of the time. CONCLUSIONS: Over the last 10 years, there has been an increase in the use of modified rotation advancement repairs and Fisher anatomic subunit approximation technique for treatment of UCL. There continues to be a lack of evidence regarding superiority of specific repair techniques or the benefits of adjunct procedures, which results in varying practice patterns. Educating all cleft surgeons on practices that are well supported is important to improve care to cleft patients.

Conservative Secondary Shell Substitution In Cyclooxygenase-2 Reduces Inhibition by Indomethacin Amides and Esters via Altered Enzyme Dynamics.

The cyclooxygenase enzymes (COX-1 and COX-2) are the therapeutic targets of nonsteroidal anti-inflammatory drugs (NSAIDs). Neutralization of the carboxylic acid moiety of the NSAID indomethacin to an ester or amide functionality confers COX-2 selectivity, but the molecular basis for this selectivity has not been completely revealed through mutagenesis studies and/or X-ray crystallographic attempts. We expressed and assayed a number of divergent secondary shell COX-2 active site mutants and found that a COX-2 to COX-1 change at position 472 (Leu in COX-2, Met in COX-1) reduced the potency of enzyme inhibition by a series of COX-2-selective indomethacin amides and esters. In contrast, the potencies of indomethacin, arylacetic acid, propionic acid, and COX-2-selective diarylheterocycle inhibitors were either unaffected or only mildly affected by this mutation. Molecular dynamics simulations revealed identical equilibrium enzyme structures around residue 472; however, calculations indicated that the L472M mutation impacted local low-frequency dynamical COX constriction site motions by stabilizing the active site entrance and slowing constriction site dynamics. Kinetic analysis of inhibitor binding is consistent with the computational findings.

Flavokawains A and B from kava (Piper methysticum) activate heat shock and antioxidant responses and protect against hydrogen peroxide-induced cell death in HepG2 hepatocytes.

Context Flavokawains are secondary metabolites from the kava plant (Piper methysticum Forst. f., Piperaceae) that have anticancer properties and demonstrated oral efficacy in murine cancer models. However, flavokawains also have suspected roles in rare cases of kava-induced hepatotoxicity. Objective To compare the toxicity flavokawains A and B (FKA, FKB) and monitor the resulting transcriptional responses and cellular adaptation in the human hepatocyte cell line, HepG2. Materials and methods HepG2 were treated with 2-100 µM FKA or FKB for 24-48 h. Cellular viability was measured with calcein-AM and changes in signalling and gene expression were monitored by luciferase reporter assay, real-time PCR and Western blot of both total and nuclear protein extracts. To test for subsequent resistance to oxidative stress, cells were pretreated with 50 µM FKA, 10 µM FKB or 10 µM sulphoraphane (SFN) for 24 h, followed by 0.4-2.8 mM H2O2 for 48 h, and then viability was assessed. Results FKA (≤100 µM) was not toxic to HepG2, whereas FKB caused significant cell death (IC50=23.2 ± 0.8 µM). Both flavokawains activated Nrf2, increasing HMOX1 and GCLC expression and enhancing total glutathione levels over 2-fold (p < 0.05). FKA and FKB also activated HSF1, increasing HSPA1A and DNAJA4 expression. Also, flavokawain pretreatment mitigated cell death after a subsequent challenge with H2O2, with FKA being more effective than FKB, and similar to SFN. Conclusions Flavokawains promote an adaptive cellular response that protects hepatocytes against oxidative stress. We propose that FKA has potential as a chemopreventative or chemotherapeutic agent.

ERK-dependent phosphorylation of HSF1 mediates chemotherapeutic resistance to benzimidazole carbamates in colorectal cancer cells.

Drugs containing the benzimidazole carbamate scaffold include anthelmintic and antifungal agents, and they are now also recognized as having potential applications in the treatment of colorectal and other cancers. These agents act by binding to ß-tubulin, and in doing so they disrupt microtubules, arrest cell division, and promote apoptotic cell death in malignant cells. We have evaluated several commercially available benzimidazole carbamates for cytotoxic activity in colorectal cancer cells. In addition to cytotoxicity, we also observe activation of the transcription factor, heat shock factor-1 (HSF1). HSF1 is well known to mediate a cytoprotective response that promotes tumor cell survival and drug resistance. Here, we show that biochemical inhibition with the HSF1 inhibitor KRIBB11 or siRNA-based silencing of HSF1 results in a significant enhancement of drug potency, causing an approximately two-fold decrease in IC50 values of parbendazole and nocodazole. We also define a mechanism for drug-induced HSF1 activation, which results from a phosphorylation event at Ser326 that is dependent on the activation of the extracellular regulated protein kinase-1/2 (ERK-1/2) mitogen-activated protein kinase pathway. Inhibition of the upstream kinase MEK-1/2 with U0126 attenuates the phosphorylation of both ERK-1/2 and HSF1, and significantly enhances drug cytotoxicity. From these data we propose a unique model whereby the ERK-1/2-dependent activation of HSF1 promotes chemotherapeutic resistance to benzimidazole carbamates. Therefore, targeting the ERK-1/2 signaling cascade is a potential strategy for HSF1 inhibition and a means of enhancing the cytotoxicity of these agents.

Fluorescent probes of the apoptolidins and their utility in cellular localization studies.

Apoptolidin A has been described among the top 0.1% most-cell-selective cytotoxic agents to be evaluated in the NCI 60 cell line panel. The molecular structure of apoptolidin A consists of a 20-membered macrolide with mono- and disaccharide moieties. In contrast to apoptolidin A, the aglycone (apoptolidinone) shows no cytotoxicity (>10 µM) when evaluated against several tumor cell lines. Apoptolidin H, the C27 deglycosylated analogue of apoptolidin A, displayed sub-micromolar activity against H292 lung carcinoma cells. Selective esterification of apoptolidins A and H with 5-azidopentanoic acid afforded azido-functionalized derivatives of potency equal to that of the parent macrolide. They also underwent strain-promoted alkyne-azido cycloaddition reactions to provide access to fluorescent and biotin-functionalized probes. Microscopy studies demonstrate apoptolidins A and H localize in the mitochondria of H292 human lung carcinoma cells.

Surgical Marking Pen Contamination: Writing a Postoperative Infection Into Your Preoperative Plan.

Introduction Preoperative marking is an essential safety practice to prevent "never" events, including wrong site surgery. Moreover, the Joint Commission regulations of the Universal Protocol require that patients be marked to indicate the operative site. Marking typically occurs with a pen or marker, which may be disposable or reusable. Previous studies have demonstrated that methicillin-resistant Staphylococcus aureus (MRSA) can survive in the dark, moist, capped environment of the marking pen and thus could plausibly be a nidus for transmission from patient to patient. The Joint Commission has established no increased risk of postoperative infection with these markings. With this study, we aimed to determine the colonization of surgical marking pens in the plastic surgery population. Methods Two marking pens from five different attending plastic surgeons at a single institution were cultured in standard fashion for aerobic and anaerobic growth. All pens were used repeatedly in office settings for performing patient markings. Those same ten marking pens were then used to mark incision sites on mock patients. Standard povidone-iodine prepping was then performed in a paint-only fashion over the skin markings, and cultures were again taken. A control group consisted of cultures from five sterile pens from the operating room. Each sterile pen was opened, uncapped, and then swabbed. All twenty-five cultures were analyzed in the hospital laboratory in a blinded fashion. Results The five control pens revealed no bacterial growth. Of the 10 direct pen cultures, two samples grew coagulase-negative staphylococci and one culture contained Pseudomonas aeruginosa. The 10-patient marked and prepped specimens showed eight negative cultures and two with coagulase-negative staphylococci. Although Pseudomonas was detected on standard pen culture, no pseudomonal growth was present in any of the samples after patient marking and prepping with povidone-iodine. Conclusions Our findings reaffirm that marking pens may be vehicles for bacterial transmission and expand upon previous studies by describing the presence of bacterial colonization on marking pens even after surgical site preparation with povidone-iodine.

Identification of PLSCR1 as a protein that interacts with RELT family members.

Receptor expressed in lymphoid tissues (RELT) proteins are recently described surface receptors belonging to the larger TNF receptor family. To improve our understanding of RELT-mediated signal transduction, we performed a screen for RELT-interacting proteins. Phospholipid Scramblase 1 (PLSCR1) was identified through a yeast two-hybrid genetic screen utilizing the intracellular portion of the RELT family member, RELL1, as bait. PLSCR1 was observed to physically interact with all known RELT family members as determined by co-immunoprecipitation experiments. The protein kinase, oxidative stress responsive 1 (OSR1) was previously shown to interact and phosphorylate all three RELT family members. In our study, no physical association was observed between OSR1 and PLSCR1 alone. However, in the presence of RELT, OSR1 was capable of co-immunoprecipitating PLSCR1, suggesting the formation of a protein complex between RELT, OSR1, and PLSCR1. In addition, OSR1 phosphorylated PLSCR1 in an in vitro kinase assay, but only in the presence of RELT, suggesting a functional multiprotein complex. RELT and PLSCR1 co-localized in intracellular regions of human embryonic kidney-293 cells, with RELT overexpression appearing to alter the localization of PLSCR1. These studies demonstrate that RELT family members physically interact with PLSCR1, and that these interactions may regulate the phosphorylation of PLSCR1 by OSR1.

Altersolanol B, a fungal tetrahydroanthraquinone, inhibits the proliferation of estrogen receptor-expressing (ER+) human breast adenocarcinoma by modulating PI3K/AKT, p38/ERK MAPK and associated signaling pathways.

The present study focused on the apoptosis-inducing effects and cellular signal-modulating properties of altersolanol B (AB), a minor fungal tetrahydroanthraquinone (THAQ) metabolite, in the estrogen receptor positive (ER+) human breast adenocarcinoma cell line, MCF-7. AB demonstrated approximately 4-fold greater antiproliferative activity in ER+ MCF-7 cells (IC50 5.5 µM) compared to the ER-negative (triple-negative) MDA-MB-231 (IC50 21.3 µM). The viability of normal breast fibrocystic epithelial cells, MCF-10A, was unaffected. AB induced intrinsic apoptosis in MCF-7 cells; it triggered the activation of caspase 9 and poly (ADP-ribose) polymerase (PARP), upregulated the expression of pro-apoptotic Bax, and downregulated the expression of anti-apoptotic Bcl-2. AB induced cell cycle arrest at G0/G1, as indicated by the downregulation of key checkpoint proteins operating at the G0/G1 phase of the cell cycle (cyclin D1, CDK4 and CDK2). The observed increase in p21Waf1/Cip1 and p53 expression may facilitate cell cycle arrest, and the subsequent induction of apoptosis. AB lacked significant effects on intracellular ROS levels, while it down-regulated nuclear factor erythroid 2-related factor 2 (Nrf2), and the Nrf2-dependent antioxidant enzyme, heme oxygenase-1. The compound disrupted AKT signaling through the downregulation of phospho-AKT and phospho-FOXO1, and the upregulation of PTEN, a phosphatase and tumor suppressor that negatively regulates the PI3K/AKT pathway. AB also disrupted the phosphorylation of AKT-controlled eukaryotic translation initiation factor, 4E-BP1, and GSK-3ß, both of which are aberrantly regulated in human cancer. The AB-dependent downregulation of NF-κB was corroborated by the inhibition of TNFα-induced NF-κB activity as monitored in a luciferase reporter. The NF-κB inhibitory activity of AB was 3-fold more potent than that of the standard inhibitor, N-p-Tosyl-l-phenylalanine chloromethyl ketone. In addition to reducing the pro-survival effects of NF-кB, the inhibition of AKT phosphorylation by AB may also lead to FOXO1-mediated growth arrest and apoptosis. AB upregulated the expression of phospho-MKK4 and phospho-p38, and downregulated the expression of phospho-MEK1/2 and phospho-ERK1/2 indicating opposing effects on the two important oncogenic signaling cascades that are aberrantly activated in many cancers. AB disrupted both the AKT and ERK1/2 signaling pathways leading to apoptosis in ER+ MCF-7 cells through mitochondria-associated mechanisms coupled with the potent inhibition of NF-кB activation. The clinical limitations of multi-agent combination therapy that targets multiple pathways in cancer may potentially be circumvented by using a single molecule, such as AB, that inhibits both AKT and ERK1/2 signaling. Our preliminary study suggested that the THAQ pharmacophore, with its disrupted conjugated ring system and relative redox inactivity, may possess greater mechanistic advantage against ER+ breast cancer when compared to the fully conjugated ring systems of the anthraquinone that possess intrinsic redox activity and DNA interacting ability. This study supports the continued investigation of THAQs as lead molecules in anticancer drug discovery and development.

A Severe Case of Siliconoma-induced Hypercalcemia due to Illicit Gluteal Silicone Injections.

Siliconoma-induced hypercalcemia is a rare complication of siliconoma, occurring secondary to a foreign body granulomatous process induced by the introduction of silicone into soft tissue. This is a case report of a woman presenting with sequelae of illicit silicone injections performed in an unknown woman's basement in Florida 20 years before presentation. A 39-year-old woman presented with a 2-month history of 20-pound weight loss, malaise, and intractable vomiting with a remote history of unregulated cosmetic injections to the bilateral gluteal and thigh regions. Her laboratory studies were consistent with severe hypercalcemia secondary to a foreign body granulomatous process. Initially, she was medically managed, with mild improvements in her symptomatic hypercalcemia and later underwent palliative debridement with siliconoma removal. Postoperatively, her course was complicated by delayed wound healing and graft failure, but the surgical defect was later closed successfully with split-thickness skin grafting after months of wound care. Although the procedure was not intended to treat her hypercalcemia, there were significant improvements in serum and ionized calcium in the months following her procedure. Severe hypercalcemia in the context of previous unregulated cosmetic injections or possible silicone implant rupture should prompt consideration of siliconoma-induced hypercalcemia as the underlying etiology. In addition to the established utility of IV fluids, bisphosphonates, and glucocorticoids, there may also be a role for surgical intervention in the management of this unique patient population's hypercalcemia.

Targeted therapy for breast cancer: An overview of drug classes and outcomes.

The last 25 years have seen significant growth in new therapeutic options for breast cancer, termed targeted therapies based on their ability to block specific pathways known to drive breast tumor growth and survival. Introduction of these drugs has been made possible through advances in the understanding of breast cancer biology. While the promise of targeted therapy for breast cancer has been clear for some time, the experience of the clinical use of multiple drugs and drug classes allows us to now present a summary and perspective as to the success and impact of this endeavor. Here we will review breast cancer targeted therapeutics in clinical use. We will provide the rationale for their indications and summarize clinical data in patients with different breast cancer subtypes, their impact on breast cancer progression and survival and their major adverse effects. The focus of this review will be on the development that has occurred within classes of targeted therapies and subsequent impact on breast cancer patient outcomes. We will conclude with a perspective on the role of targeted therapy in breast cancer treatment and highlight future areas of development.

Lower Extremity Soft Tissue Reconstruction Review Article.

Reconstruction plays a valuable role in the management of lower extremity wounds for limb salvage. The goals of reconstruction are to improve function and quality of life, return to work, and pain reduction while providing a long-lasting durable reconstruction. The plastics and reconstructive surgical approach in conjunction with the orthopedic or trauma team, referred often as the "orthoplastic" approach, can yield the best outcomes for patients. The following sections discuss reconstruction principles and techniques that can be applied broadly for lower extremity wounds secondary to trauma, infection, and tumor resection.

Long patch base excision repair compensates for DNA polymerase ß inactivation by the C4'-oxidized abasic site.

The C4'-oxidized abasic site (C4-AP), which is produced by a variety of damaging agents, has significant consequences for DNA. The lesion is highly mutagenic and reactive, resulting in interstrand cross-links. The base excision repair of DNA containing independently generated C4-AP was examined. C4-AP is incised by Ape1 ~12-fold less efficiently than an apurinic/apyrimidinic lesion. DNA polymerase ß induces the ß-elimination of incised C4-AP in ternary complexes, duplexes, and single-stranded substrate. However, excision from a ternary complex is most rapid. In addition, the lesion inactivates the enzyme after approximately seven turnovers on average by reacting with one or more lysine residues in the lyase active site. Unlike 5'-(2-phosphoryl-1,4-dioxobutane), which very efficiently irreversibly inhibits DNA polymerase ß, the lesion is readily removed by strand displacement synthesis conducted by the polymerase in conjunction with flap endonuclease 1. DNA repair inhibition by C4-AP may be a partial cause of the cytotoxicity of drugs that produce this lesion.

Product and mechanistic analysis of the reactivity of a C6-pyrimidine radical in RNA.

Nucleobase radicals are the major reactive intermediates produced when hydroxyl radical reacts with nucleic acids. 5,6-Dihydrouridin-6-yl radical (1) was independently generated from a ketone precursor via Norrish Type I photocleavage in a dinucleotide, single-stranded, and double-stranded RNA. This radical is a model of the major hydroxyl radical adduct of uridine. Tandem lesions resulting from addition of the peroxyl radical derived from 1 to the 5'-adjacent nucleotide are observed by ESI-MS. Radical 1 produces direct strand breaks at the 5'-adjacent nucleotide and at the initial site of generation. The preference for cleavage at these two positions depends upon the secondary structure of the RNA and whether O(2) is present or not. Varying the identity of the 5'-adjacent nucleotide has little effect on strand scission. In general, strand scission is significantly more efficient under anaerobic conditions than when O(2) is present. Strand scission is more than twice as efficient in double-stranded RNA than in a single-stranded oligonucleotide under anaerobic conditions. Internucleotidyl strand scission occurs via ß-fragmentation following C2'-hydrogen atom abstraction by 1. The subsequently formed olefin cation radical ultimately yields products containing 3'-phosphate or 3'-deoxy-2'-ketouridine termini. These end groups are proposed to result from competing deprotonation pathways. The dependence of strand scission efficiency from 1 on secondary structure under anaerobic conditions suggests that this reactivity may be useful for extracting additional RNA structural information from hydroxyl radical reactions.

Systems analysis of protein modification and cellular responses induced by electrophile stress.

Biological electrophiles result from oxidative metabolism of exogenous compounds or endogenous cellular constituents, and they contribute to pathophysiologies such as toxicity and carcinogenicity. The chemical toxicology of electrophiles is dominated by covalent addition to intracellular nucleophiles. Reaction with DNA leads to the production of adducts that block replication or induce mutations. The chemistry and biology of electrophile-DNA reactions have been extensively studied, providing in many cases a detailed understanding of the relation between adduct structure and mutational consequences. By contrast, the linkage between protein modification and cellular response is poorly understood. In this Account, we describe our efforts to define the chemistry of protein modification and its biological consequences using lipid-derived alpha,beta-unsaturated aldehydes as model electrophiles. In our global approach, two large data sets are analyzed: one represents the identity of proteins modified over a wide range of electrophile concentrations, and the second comprises changes in gene expression observed under similar conditions. Informatics tools show theoretical connections based primarily on transcription factors hypothetically shared between the two data sets, downstream of adducted proteins and upstream of affected genes. This method highlights potential electrophile-sensitive signaling pathways and transcriptional processes for further evaluation. Peroxidation of cellular phospholipids generates a complex mixture of both membrane-bound and diffusible electrophiles. The latter include reactive species such as malondialdehyde, 4-oxononenal, and 4-hydroxynonenal (HNE). Enriching HNE-adducted proteins for proteomic analysis was a technical challenge, solved with click chemistry that generated biotin-tagged protein adducts. For this purpose, HNE analogues bearing terminal azide or alkyne functionalities were synthesized. Cellular lysates were first exposed to a single type of HNE analogue (azido- or alkynyl-HNE), and then click reactions were performed against the cognate alkynyl- and azido-biotin derivative. The resulting biotin-labeled proteins were captured and enriched over a streptavidin matrix for subsequent mass spectrometric analysis. We thereby identified a multitude of HNE targets. Simultaneous microarray analysis of changes in gene expression triggered by HNE also produced an abundance of data. Functional analysis of both data sets generated the hypothesis that an important pathway of cellular response derives from electrophile modification of protein chaperones, resulting in the release of transcription factors that are their clients. Informatic analysis of the protein modification and microarray data sets identified several transcription factors as potential mediators of the cellular response to HNE-adducted proteins. Among these, heat shock factor 1 (HSF1) was confirmed as a sensitive and robust effector of HNE-induced changes in gene expression. Activation of HSF1 appears, in part, to be mediated by the electrophilic adduction of Hsp70 and Hsp90, which normally maintain HSF1 in an inactive cytosolic complex. The identification of HSF1 as a mediator of biological effects downstream of HSF1 has provided new opportunities for research, illustrating the potential of our systems-based approach. Accordingly, we characterized HSF1-mediated gene expression in protecting against electrophile-induced toxicity. Among the genes induced by HSF1, Bcl-2- associated athanogene 3 (BAG3) is notable for its actions in promoting cell survival through stabilization of antiapoptotic Bcl-2 proteins, appearing to have a critical role in mediating cellular protection against electrophile-induced death.

Direct strand scission from a nucleobase radical in RNA.

RNA oxidation is important in the etiology of disease and as a tool for studying the structure and folding kinetics of this biopolymer. Nucleobase radicals are the major family of reactive intermediates produced in RNA exposed to diffusible species such as hydroxyl radical. The nucleobase radicals are believed to produce direct strand breaks by abstracting hydrogen atoms from their own and neighboring ribose rings. By independently generating the formal C5 hydrogen atom addition product of uridine in RNA, we provide the first chemical characterization of the pathway for direct strand scission from an RNA nucleobase radical. The process is more efficient under anaerobic conditions. The preference for strand scission in double-stranded RNA over single-stranded RNA suggests that this chemistry may be useful for analyzing the secondary structure of RNA in hydroxyl radical cleavage experiments if they are carried out under anaerobic conditions.

Orthostatic proteinuria and the spectrum of diurnal variability of urinary protein excretion in healthy children.

The aim of this study was to characterize the 24-h and diurnal variability of urinary protein excretion and identify the prevalence of orthostatic proteinuria (OP) in healthy children. Upright, supine, and 24-h total urinary protein (UrTP) and creatinine clearance (CrCl) were measured in 91 healthy children ages 6-19 years. Urinary protein and creatinine excretions were calculated and examined by gender, age, Tanner stage, and body mass index (BMI). Orthostatic proteinuria (OP) was defined as a 24-h UrTP >100 mg/m(2) with a normal supine UrTP (<4 mg/m(2)/h). There exists a marked diurnal variability in UrTP. The upright UrTP rate was three to four-times greater than the supine rate. UrTP, adjusted for body surface area, is higher in boys than girls and increases with age and BMI. There is a similar increase in upright CrCl compared with supine. Urinary protein to creatinine ratio (UPcr) is strongly correlated with UrTP. OP is common, being found in 20% of children in this cohort, and is more common in boys and associated with age >10 years and BMI >85%. In children with OP, a first morning UPcr shows a value in the normal range, whereas a random daytime UPcr is elevated. There exists a diurnal variability in urinary protein excretion that is exaggerated in participants with OP. UPcr reliably estimates 24-h UrTP. Using current pediatric criteria, OP is very common, particularly in boys. A normal first morning UPcr ratio indicates that a child with elevated random urinary protein has OP.

Breast Adipocyte Co-culture Increases the Expression of Pro-angiogenic Factors in Macrophages.

Obese individuals with breast cancer have a poorer prognosis and higher risk of metastatic disease vs. non-obese patients. Adipose tissue in obese individuals is characterized by an enhanced macrophage infiltration, creating a microenvironment that favors tumor progression. Here, we demonstrate a role for adipocyte-macrophage interactions in the regulation of angiogenesis. Co-culture of THP-1 macrophages with human breast adipocytes led to increased expression of the pro-angiogenic growth factor, vascular endothelial growth factor A (VEGFA). Several adipocyte-derived proteins including leptin, insulin, IL-6, and TNF-α were each capable of increasing VEGFA expression in THP-1 macrophages, identifying these as possible mediators of the changes that were observed with co-culture. Furthermore, analysis of THP-1 culture media by antibody array revealed that THP-1 secrete several other pro-angiogenic signals in response to adipocyte co-culture, including interleukin 8 (IL-8), matrix metalloproteinase 9 (MMP9), pentraxin 3 (PTX3), and serpin E1 (plasminogen activator inhibitor 1, PAI1) after co-culture with human adipocytes. We used an in vitro endothelial tube formation assay with human vascular endothelial cells to evaluate the effects of THP-1 culture media on angiogenesis. Here, culture media from THP-1 cells previously exposed to human adipocytes stimulated endothelial tube formation more significantly than THP-1 cells cultured alone. In summary, we find that adipocyte co-culture stimulates the expression of pro-angiogenic mediators in macrophages and has pro-angiogenic effects in vitro, thus representing a possible mechanism for the enhanced risk of breast cancer progression in obese individuals.

RELT stains prominently in B-cell lymphomas and binds the hematopoietic transcription factor MDFIC.

Receptor Expressed in Lymphoid Tissues (RELT) is a human tumor necrosis factor receptor superfamily member (TNFRSF) that is expressed most prominently in cells and tissues of the hematopoietic system. RELL1 and RELL2 are two homologs that physically interact with RELT and co-localize with RELT at the plasma membrane. This study sought to further elucidate the function of RELT by identifying novel protein interactions with RELT family members. The transcription factor MyoD family inhibitor domain-containing (MDFIC) was identified in a yeast two-hybrid genetic screen using RELL1 as bait. MDFIC co-localizes with RELT family members at the plasma membrane; this co-localization was most prominently observed with RELL1 and RELL2. In vitro co-immunoprecipitation (Co-IP) was utilized to demonstrate that MDFIC physically interacts with RELT, RELL1, and RELL2. Co-IP using deletion mutants of MDFIC and RELT identified regions important for physical association between MDFIC and RELT family members and a computational analysis revealed that RELT family members are highly disordered proteins. Immunohistochemistry of normal human lymph nodes revealed RELT staining that was most prominent in macrophages. Interestingly, the level of RELT staining significantly increased progressively in low and high-grade B-cell lymphomas versus normal lymph nodes. RELT co-staining with CD20 was observed in B-cell lymphomas, indicating that RELT is expressed in malignant B cells. Collectively, these results further our understanding of RELT-associated signaling pathways, the protein structure of RELT family members, and provide preliminary evidence indicating an association of RELT with B-cell lymphomas.