Effect of a Low-Protein Diet Supplemented with Ketoacids on Skeletal Muscle Atrophy and Autophagy in Rats with Type 2 Diabetic Nephropathy

A low-protein diet supplemented with ketoacids maintains nutritional status in patients with diabetic nephropathy. The activation of autophagy has been shown in the skeletal muscle of diabetic and uremic rats. This study aimed to determine whether a low-protein diet supplemented with ketoacids improves muscle atrophy and decreases the increased autophagy observed in rats with type 2 diabetic nephropathy. In this study, 24-week-old Goto-Kakizaki male rats were randomly divided into groups that received either a normal protein diet (NPD group), a low-protein diet (LPD group) or a low-protein diet supplemented with ketoacids (LPD+KA group) for 24 weeks. Age- and weight-matched Wistar rats served as control animals and received a normal protein diet (control group). We found that protein restriction attenuated proteinuria and decreased blood urea nitrogen and serum creatinine levels. Compared with the NPD and LPD groups, the LPD+KA group showed a delay in body weight loss, an attenuation in soleus muscle mass loss and a decrease of the mean cross-sectional area of soleus muscle fibers. The mRNA and protein expression of autophagy-related genes, such as Beclin-1, LC3B, Bnip3, p62 and Cathepsin L, were increased in the soleus muscle of GK rats fed with NPD compared to Wistar rats. Importantly, LPD resulted in a slight reduction in the expression of autophagy-related genes; however, these differences were not statistically significant. In addition, LPD+KA abolished the upregulation of autophagy-related gene expression. Furthermore, the activation of autophagy in the NPD and LPD groups was confirmed by the appearance of autophagosomes or autolysosomes using electron microscopy, when compared with the Control and LPD+KA groups. Our results showed that LPD+KA abolished the activation of autophagy in skeletal muscle and decreased muscle loss in rats with type 2 diabetic nephropathy.     

Caspase-2 Short Isoform Interacts with Membrane-Associated Cytoskeleton Proteins to Inhibit Apoptosis

Caspase-2 (casp-2) is the most conserved caspase across species, and is one of the initiator caspases activated by various stimuli. The casp-2 gene produces several alternative splicing isoforms. It is believed that the long isoform, casp-2L, promotes apoptosis, whereas the short isoform, casp-2S, inhibits apoptosis. The actual effect of casp-2S on apoptosis is still controversial, however, and the underlying mechanism for casp-2S-mediated apoptosis inhibition is unclear. Here, we analyzed the effects of casp-2S on DNA damage induced apoptosis through “gain-of-function” and “loss-of-function” strategies in ovarian cancer cell lines. We clearly demonstrated that the over-expression of casp-2S inhibited, and the knockdown of casp-2S promoted, the cisplatin-induced apoptosis of ovarian cancer cells. To explore the mechanism by which casp-2S mediates apoptosis inhibition, we analyzed the proteins which interact with casp-2S in cells by using immunoprecipitation (IP) and mass spectrometry. We have identified two cytoskeleton proteins, Fodrin and α-Actinin 4, which interact with FLAG-tagged casp-2S in HeLa cells and confirmed this interaction through reciprocal IP. We further demonstrated that casp-2S (i) is responsible for inhibiting DNA damage-induced cytoplasmic Fodrin cleavage independent of cellular p53 status, and (ii) prevents cisplatin-induced membrane blebbing. Taken together, our data suggests that casp-2S affects cellular apoptosis through its interaction with membrane-associated cytoskeletal Fodrin protein.     

Identification and design of a novel series of MGAT2 inhibitors

[Acyl CoA]monoacylglycerol acyltransferase 2 (MGAT2) is of interest as a target for therapeutic treatment of diabetes, obesity and other diseases which together constitute the metabolic syndrome. In this Letter we report our discovery and optimisation of a novel series of MGAT2 inhibitors. The development of the SAR of the series and a detailed discussion around some key parameters monitored and addressed during the lead generation phase will be given. The in vivo results from an oral lipid tolerance test (OLTT) using the MGAT2 inhibitor (S)-10, shows a significant reduction (68% inhibition relative to na?ve, p <0.01) in plasma triacylglycerol (TAG) concentration.     

Glioma IL13Rα2 Is Associated with Mesenchymal Signature Gene Expression and Poor Patient Prognosis

A major challenge for successful immunotherapy against glioma is the identification and characterization of validated targets. We have taken a bioinformatics approach towards understanding the biological context of IL-13 receptor α2 (IL13Rα2) expression in brain tumors, and its functional significance for patient survival. Querying multiple gene expression databases, we show that IL13Rα2 expression increases with glioma malignancy grade, and expression for high-grade tumors is bimodal, with approximately 58% of WHO grade IV gliomas over-expressing this receptor. By several measures, IL13Rα2 expression in patient samples and low-passage primary glioma lines most consistently correlates with the expression of signature genes defining mesenchymal subclass tumors and negatively correlates with proneural signature genes as defined by two studies. Positive associations were also noted with proliferative signature genes, whereas no consistent associations were found with either classical or neural signature genes. Probing the potential functional consequences of this mesenchymal association through IPA analysis suggests that IL13Rα2 expression is associated with activation of proinflammatory and immune pathways characteristic of mesenchymal subclass tumors. In addition, survival analyses indicate that IL13Rα2 over-expression is associated with poor patient prognosis, a single gene correlation ranking IL13Rα2 in the top ~1% of total gene expression probes with regard to survival association with WHO IV gliomas. This study better defines the functional consequences of IL13Rα2 expression by demonstrating association with mesenchymal signature gene expression and poor patient prognosis. It thus highlights the utility of IL13Rα2 as a therapeutic target, and helps define patient populations most likely to respond to immunotherapy in present and future clinical trials.     

Prospective study of bacterial meningitis in North East Thames region, 1991-3, during introduction of Haemophilus influenzae vaccine.

OBJECTIVE--To describe the epidemiology of primary bacterial meningitis in the North East Thames region over a three year period before and during the introduction of the vaccine for Haemophilus influenzae type b. DESIGN--Analysis of information on cases of primary bacterial meningitis identified by microbiology laboratories in the region, with collection of case data by questionnaire. MAIN OUTCOME MEASURES--Annual incidence rates for types of meningitis according to age and ethnic group. RESULTS--The annual incidence rates for the three major causes of bacterial meningitis in the general population were 1.9/100,000 for Neisseria meningitidis, 1.6/100,000 for Haemophilus influenzae before vaccination, and 1.0/100,000 for Streptococcus pneumoniae. Higher rates of H influenzae meningitis were found in Asians compared with white people (3.6/100,000 v 1.5/100,000, P = 0.01). As a result of the vaccine programme introduced in October 1992 the number of cases of H influenzae meningitis in children under 5 years has fallen by 87%. CONCLUSIONS--Bacterial meningitis is a serious problem especially in preschool children. There are differences in the incidence of some causes of bacterial meningitis in different ethnic groups; with H influenzae type b being significantly more common among black and Asian people than among white people. The immunisation programme for H influenzae type b in the North East Thames region has been successful in reducing the incidence of this type of meningitis in Asian and white populations. The numbers were too small to evaluate in the black population.     

Denaturing Reversed-Phase HPLC Using a Mobile Phase Containing Urea for Oligodeoxynucleotide Analysis

Denaturing reversed-phase (RP) high performance liquid chromatography (HPLC) is usually achieved by elevating column temperature. In this article, an alternative method involving using a mobile phase that contains urea and performing HPLC at room temperature is described. The efficacy of the new method was demonstrated by analyzing a 61-mer oligodeoxynucleotide (ODN) and double-stranded (ds) ODNs. The multiple peaks of the 61-mer ODN under normal conditions merged into one under the denaturing conditions. The broad single peaks of dsODNs under normal conditions were split into two sharp peaks.     

Tampering with cancer chemoresistance by targeting the TGM2-IL6-autophagy regulatory network

Macroautophagy/autophagy is a well-established process involved in maintaining cellular homeostasis, but its role in cancer is complex and even controversial. Many studies have reported a correlative relationship between increased autophagy and evolving cancer cells under stress conditions such as nutrient or oxygen deprivation; however, there has been a lack of a plausible mechanistic link to properly target the autophagy process in the context of this microenvironment. We recently unveiled a positive regulatory loop involving TGM2 (transglutaminase 2)-NFKB/NF-κB signaling, IL6 and autophagy in cancer using mantle cell lymphoma (MCL) as a model system. These pathways are functionally connected to each other, thereby promoting malignant B cell survival and leading to enhanced lymphoma progression both in mice and in patients. Disruption of this network could provide an opportunity to increase the efficacies of current therapies and to reduce MCL drug resistance.