Erk5 is a mediator to TGFβ1-induced loss of phenotype and function in human podocytes.

Background: Podocytes are highly specialized cells integral to the normal functioning kidney, however, in diabetic nephropathy injury occurs leading to a compromised phenotype and podocyte dysfunction which critically produces podocyte loss with subsequent renal impairment. TGFβ1 holds a major role in the development of diabetic nephropathy. Erk5 is an atypical mitogen-activated protein (MAP) kinase involved in pathways modulating cell survival, proliferation, differentiation, and motility. Accordingly, the role of Erk5 in mediating TGFβ1-induced podocyte damage was investigated. Methods: Conditionally immortalized human podocytes were stimulated with TGFβ1 (2.5 ng/ml); inhibition of Erk5 activation was conducted with the chemical inhibitor BIX02188 (10 μM) directed to the upstream Mek5; inhibition of Alk5 was performed with SB431542 (10 μM); Ras signaling was inhibited with farnesylthiosalicylic acid (10 μM). Intracellular signaling proteins were investigated by western blotting; phenotype was explored by immunofluorescence; proliferation was assessed with a MTS assay; motility was examined with a scratch assay; barrier function was studied using electric cell-substrate impedance sensing; apoptosis was studied with annexin V-FITC flow cytometry. Results: Podocytes expressed Erk5 which was phosphorylated by TGFβ1 via Mek5, whilst not involving Ras. TGFβ1 altered podocyte phenotype by decreasing P-cadherin staining and increasing α-SMA, as well as reducing podocyte barrier function; both were prevented by inhibiting Erk5 phosphorylation with BIX02188. TGFβ1-induced podocyte proliferation was prevented by BIX02188, whereas the induced apoptosis was not. Podocyte motility was reduced by BIX02188 alone and further diminished with TGFβ1 co-incubation. Conclusion: These results describe for the first time the expression of Erk5 in podocytes and identify it as a potential target for the treatment of diabetic renal disease

Functional multisite copolymer by one-pot sequential RAFT copolymerization of styrene and maleic anhydride

A Multisite copolymer with functionalizable units inserted at precise locations was synthesised by one-pot Reversible Addition–Fragmentation Chain-Transfer (RAFT) polymerization and sequential Single Monomer Unit Insertion (SMUI) and Chain Extension (ChainExt) using Styrene (Sty) and Maleic Anhydride (MAnh) as comonomers. The multisite copolymer was based on a polystyrene (PSty) backbone (ca. 5700 g mol−1) with MAnh units inserted locally at four positions in the backbone. First, a well-defined macroCTA (1400 g mol−1 – Đ = 1.07) was synthesised by optimized RAFT polymerization (high conversion, high livingness and low dispersity) of styrene (DP = 10) using industrial grade butyl-2-methyl-2-[(dodecylsulfanylthiocarbonyl)sulfanyl] propionate as chain transfer agent (CTA-Ester – 80% pure). Subsequently, the polystyrene macroCTA was used for one-pot SMUI using a small excess of MAnh monomer (DPtarget = 1.5). The copolymer was chain extended by styrene leading to a polystyrene backbone with MAnh units (1.5 in average) located in the middle of the chain. By repeating SMUI and ChainExt, several units of MAnh were inserted locally along the polystyrene backbone (every 10 units on average) to give a functionalizable multisite copolymer (Đ = 1.35). Long alkyl chains (stearyl) were added by esterification of maleic anhydride moieties to obtain branched architecture

Symptoms of Gastro-Oesophageal Reflux Disease and the Severity of Obstructive Sleep Apnoea Syndrome Are Not Related in Sleep Disorders Center Patients

Background: Studies suggest obstructive sleep apnea syndrome (OSAS) frequently manifests in patients with gastroesophageal reflux disease (GERD) and that there may be a causal relationship. Aim: To determine the relationship between OSAS and symptoms of GERD. Methods: Consecutive patients referred to the Sleep Disorders Center (SDC) 18 years and older with polysomnographically defined OSAS were evaluated prospectively for GERD using a validated symptoms questionnaire. The GERD and OSAS relationship was assessed by 1) determining frequency of GERD in patients with and without OSAS; 2) ascertaining the relationship between OSAS severity categories and presence of GERD; 3) examining GERD score in relation to those factors that might affect both GERD and OSAS, e.g. obesity. Results: One thousand and twenty-three SDC patients met entry citeria. Amongst participants, GERD was common (29% of women and 17% of males) and OSAS extremely common (58% of women and 80% of males). GERD score did not correlate with OSAS variables. The severity of OSAS did not influence the prevalence of GERD. Conclusion: In a large group of patients referred to a sleep disorders center, there was no relationship between OSAS and GERD symptoms. Also, there was no relationship between the severity of OSAS and the likelihood of GERD symptoms

The structure of the ternary Eg5–ADP–ispinesib complex

The human kinesin Eg5 is responsible for bipolar spindle formation during early mitosis. Inhibition of Eg5 triggers the formation of monoastral spindles, leading to mitotic arrest that eventually causes apoptosis. There is increasing evidence that Eg5 constitutes a potential drug target for the development of cancer chemotherapeutics. The most advanced Eg5-targeting agent is ispinesib, which exhibits potent antitumour activity and is currently in multiple phase II clinical trials. In this study, the crystal structure of the Eg5 motor domain in complex with ispinesib, supported by kinetic and thermodynamic binding data, is reported. Ispinesib occupies the same induced-fit pocket in Eg5 as other allosteric inhibitors, making extensive hydrophobic interactions with the protein. The data for the Eg5-ADP-ispinesib complex suffered from pseudo-merohedral twinning and revealed translational noncrystallographic symmetry, leading to challenges in data processing, space-group assignment and structure solution as well as in refinement. These complications may explain the lack of available structural information for this important agent and its analogues. The present structure represents the best interpretation of these data based on extensive data-reduction, structure-solution and refinement trials

Therapeutic target-site variability in α1-antitrypsin characterized at high resolution

The intrinsic propensity of [alpha]1-antitrypsin to undergo conformational transitions from its metastable native state to hyperstable forms provides a motive force for its antiprotease function. However, aberrant conformational change can also occur via an intermolecular linkage that results in polymerization. This has both loss-of-function and gain-of-function effects that lead to deficiency of the protein in human circulation, emphysema and hepatic cirrhosis. One of the most promising therapeutic strategies being developed to treat this disease targets small molecules to an allosteric site in the [alpha]1-antitrypsin molecule. Partial filling of this site impedes polymerization without abolishing function. Drug development can be improved by optimizing data on the structure and dynamics of this site. A new 1.8 Å resolution structure of [alpha]1-antitrypsin demonstrates structural variability within this site, with associated fluctuations in its upper and lower entrance grooves and ligand-binding characteristics around the innermost stable enclosed hydrophobic recess. These data will allow a broader selection of chemotypes and derivatives to be tested in silico and in vitro when screening and developing compounds to modulate conformational change to block the pathological mechanism while preserving function

Challenges and surprises that arise with nucleic acids during model building and refinement

The challenges that arise in nucleic acid model building as a consequence of their simpler and more symmetric super-secondary structures are addressed

BamA β16C strand and periplasmic turns are critical for outer membrane protein insertion and assembly

Outer membrane β-barrel proteins play important roles in importing nutrients, exporting wastes and conducting signals in Gram-negative bacteria, mitochondria and chloroplasts. The outer membrane proteins are inserted and assembled into the outer membrane by OMP85 family proteins. In Escherichia coli , the b-barrel assembly machinery (BAM) contains four lipoproteins BamB, BamC, BamD and BamE, and one outer membrane protein BamA, forming a "top hat"-like structure. Structural and functional studies of the E. coli BAM machinery have revealed that the rotation of periplasmic ring may trigger the barrel b1C-b6C scissor-like movement that promote the unfolded outer membrane protein insertion without using ATP. Here we report the BamA C-terminal barrel structure of Salmonella enterica Typhimurium str. LT2 and functional assays, which reveal that the BamA's C-terminal residue Trp, the b16C strand of the barrel and the periplasmic turns are critical for the functionality of BamA. These findings indicate that the unique b16C and the periplasmic turns of BamA are important for the out membrane insertion and assembly. The periplasmic turns might mediate the rotation of the periplasmic ring to the scissor-like movement of BamA b1C-b6C, triggering the outer membrane protein insertion. These results are important for understanding the outer membrane protein insertion in Gram-negative bacteria, as well as in mitochondria and chloroplasts

A lower bound on the local extragalactic magnetic field

Assuming that the hard gamma-ray emission of Cen A is a result of synchrotron radiation of ultra-relativistic electrons, we derive a lower bound on the local extragalactic magnetic field, $B> 10^{-8}$ G. This result is consistent with (and close to) upper bounds on magnetic fields derived from consideration of cosmic microwave background distortions and Faraday rotation measurements.Comment: Includes extensive discussion of particle acceleration above 10^20 eV in the hot spot-like region of Cen

Lung Cancer Screening and Epigenetics in African Americans: The Role of the Socioecological Framework

Lung cancer is the leading cause of cancer morbidity and mortality in the U.S. and racial/ethnic minorities carry the greatest burden of lung cancer disparities with African Americans (AAs) impacted disproportionately. Inequities in lung cancer health disparities are often associated with multiple bio-behavioral and socio-cultural factors among racial/ethnic minorities. Epigenetic research has advanced the understanding of the intersectionality between biological and socio-cultural factors in lung cancer disparities among AAs. However, gaps exist in the engagement of diverse populations in epigenetic lung cancer research, which poses a challenge in ensuring the generalizability and implementation of epigenetic research in populations that carry an unequal cancer burden. Grounding epigenetic lung cancer research within a socio-ecological framework may prove promising in implementing a multi-level approach to community engagement, screening, navigation, and research participation among AAs. The University of Illinois Cancer Center (UI Cancer Center) is employing an evidence–based (EB) model of community/patient engagement utilizing the socio-ecological model (SEM) to develop a culturally sensitive epigenetic lung cancer research program that addresses multiple factors that impact lung cancer outcomes in AAs. By implementing epigenetic research within a group of Federally Qualified Health Centers (FQHCs) guided by the SEM, the UI Cancer Center is proposing a new pathway in mitigating lung cancer disparities in underserved communities. At the individual level, the framework examines tobacco use among patients at FQHCs (the organizational level) and also tailors epigenetic research to explore innovative biomarkers in high risk populations. Interpersonal interventions use Patient Navigators to support navigation to EB tobacco cessation resources and lung cancer screening. Community level support within the SEM is developed by ongoing partnerships with local and national partners such as the American Lung Association (ALA) and the American Cancer Society (ACS). Lastly, at the policy level, the UI Cancer Center acknowledges the role of policy implications in lung cancer screening and advocates for policies and screening recommendations that examine the current guidelines from the United States Preventive Services Task Force (USPTF)