A CRYGC gene mutation associated with autosomal dominant pulverulent cataract

Purpose

To describe at molecular level a family with pulverulent congenital cataract associated with a CRYGC gene mutation.

Methods

One family with several affected members with pulverulent congenital cataract and 230 healthy controls were examined. Genomic DNA from leukocytes was isolated to analyze the CRYGA-D cluster, CX46, CX50 and MIP genes through high-resolution melting curve and DNA sequencing.

Results

DNA sequencing in the affected members revealed the c.143G>A mutation (p.R48H) in exon 2 of the CRYGC gene; 230 healthy controls and ten healthy relatives were also analyzed and none of them showed the c.143G>A mutation. No other polymorphisms or mutations were found to be present.

Conclusion

In the present study, we described a family with pulverulent congenital cataract that segregated the c.143G>A mutation (p.R48H) in the CRYGC gene. A few mutations have been described in the CRYGC gene in autosomal dominant cataract, none of them with pulverulent cataract making clear the clinical heterogeneity of congenital cataract. This mutation has been associated with the phenotype of congenital cataract but also is considered an SNP in the NCBI data base. Our data and previous report suggest that p.R48H could be a disease-causing mutation and not an SNP.     

In-silico drug screening and potential target identification for hepatocellular carcinoma using Support Vector Machines based on drug screening result

Hepatocellular carcinoma (HCC) is a severe liver malignancy with few drug treatment options. In finding an effective treatment for HCC, screening drugs that are already FDA-approved will fast track the clinical trial and drug approval process. Connectivity Map (CMap), a large repository of chemical-induced gene expression profiles, provides the opportunity to analyze drug properties on the basis of gene expression. Support Vector Machines (SVM) were utilized to classify the effectiveness of drugs against HCC using gene expression profiles in CMap. The results of this classification will help us (1) identify genes that are chemically sensitive, and (2) predict the effectiveness of remaining chemicals in CMap in the treatment of HCC and provide a prioritized list of possible HCC drugs for biological verification.     

Ezrin-Radixin-Moesin-binding Phosphoprotein 50 (EBP50) and Nuclear Factor-κB (NF-κB): A FEED-FORWARD LOOP FOR SYSTEMIC AND VASCULAR INFLAMMATION*

The interaction between vascular cells and macrophages is critical during vascular remodeling. Here we report that the scaffolding protein, ezrin-binding phosphoprotein 50 (EBP50), is a central regulator of macrophage and vascular smooth muscle cells (VSMC) function. EBP50 is up-regulated in intimal VSMC following endoluminal injury and promotes neointima formation. However, the mechanisms underlying these effects are not fully understood. Because of the fundamental role that inflammation plays in vascular diseases, we hypothesized that EBP50 mediates macrophage activation and the response of vessels to inflammation. Indeed, EBP50 expression increased in primary macrophages and VSMC, and in the aorta of mice, upon treatment with LPS or TNFα. This increase was nuclear factor-κB (NF-κB)-dependent. Conversely, activation of NF-κB was impaired in EBP50-null VSMC and macrophages. We found that inflammatory stimuli promote the formation of an EBP50-PKCζ complex at the cell membrane that induces NF-κB signaling. Macrophage activation and vascular inflammation after acute LPS treatment were reduced in EBP50-null cells and mice as compared with WT. Furthermore, macrophage recruitment to vascular lesions was significantly reduced in EBP50 knock-out mice. Thus, EBP50 and NF-κB participate in a feed-forward loop leading to increased macrophage activation and enhanced response of vascular cells to inflammation.     

(−)-Reboxetine inhibits muscle nicotinic acetylcholine receptors by interacting with luminal and non-luminal sites

The interaction of (−)-reboxetine, a non-tricyclic norepinephrine selective reuptake inhibitor, with muscle-type nicotinic acetylcholine receptors (AChRs) in different conformational states was studied by functional and structural approaches. The results established that (−)-reboxetine: (a) inhibits (±)-epibatidine-induced Ca²⁺ influx in human (h) muscle embryonic (hα1β1γδ) and adult (hα1β1εδ) AChRs in a non-competitive manner and with potencies IC₅₀ = 3.86 ± 0.49 and 1.92 ± 0.48 μM, respectively, (b) binds to the [³H]TCP site with ∼13-fold higher affinity when the Torpedo AChR is in the desensitized state compared to the resting state, (c) enhances [³H]cytisine binding to the resting but activatableTorpedo AChR but not to the desensitized AChR, suggesting desensitizing properties, (d) overlaps the PCP luminal site located between rings 6′ and 13′ in the Torpedo but not human muscle AChRs. In silico mutation results indicate that ring 9′ is the minimum structural component for (−)-reboxetine binding, and (e) interacts to non-luminal sites located within the transmembrane segments from the Torpedo AChR γ subunit, and at the α1/ε transmembrane interface from the adult muscle AChR. In conclusion, (−)-reboxetine non-competitively inhibits muscle AChRs by binding to the TCP luminal site and by inducing receptor desensitization (maybe by interacting with non-luminal sites), a mechanism that is shared by tricyclic antidepressants.     

A metabonomic investigation of the effects of 60 days exposure of rats to two types of pyrethroid insecticides

Type I and II pyrethroid insecticides display different neurotoxicity. To investigate the long-term (60 days exposure) metabolic effect of the two types of pyrethroid insecticides deltamethrin and permethrin, ¹H nuclear magnetic resonance (NMR) spectroscopy-based metabonomics was used to analyze the biochemical composition of urine and serum samples from rats administrated daily with deltamethrin or permethrin for 60 consecutive days, and principal component analysis used to visualize similarities and differences in the resultant biochemical profiles. Rats treated with either deltamethrin or permethrin displayed increased levels of urinary acetate, dimethylamine, dimethylglycine, trimethylamine and serum free amino acids, and decreased urinary 2-oxoglutarate, all of which are indicative of kidney lesions and nephrotoxicity. The reduced excretion of tricarboxylic acid cycle intermediates, together with increased 3-D-hydroxybutyrate, acetate, and lactate in treated rats could suggest disturbance of the energy metabolism, including an increased rate of anaerobic glycolysis, enhanced fatty acid β-oxidation and ketogenesis. These results show that these two types of insecticides have similarities in the urine and serum spectra, indicating that similar metabolic pathways are perturbed by the insecticides, which induced hepatotoxicity and nephrotoxicity. This approach may lead to the discovery of novel biomarkers of pyrethroids toxicity and thereby provide new insights into the toxicological mechanisms of pesticides pyrethroids.     

Bioavailability and potential carcinogenicity of polycyclic aromatic hydrocarbons from wood combustion particulate matter in vitro

Due to increasing energy demand and limited fossil fuels, renewable energy sources have gained in importance. Particulate matter (PM) in general, but also PM from the combustion of wood is known to exert adverse health effects in human. These are often related to specific toxic compounds adsorbed to the PM surface, such as polycyclic aromatic hydrocarbons (PAH), of which some are known human carcinogens. This study focused on the bioavailability of PAHs and on the tumor initiation potential of wood combustion PM, using the PAH CALUX® reporter gene assay and the BALB/c 3T3 cell transformation assay, respectively. For this, both cell assays were exposed to PM and their respective organic extracts from varying degrees of combustion. The PAH CALUX® experiments demonstrated a concentration–response relationship matching the PAHs detected in the samples. Contrary to expectations, PM samples from complete (CC) and incomplete combustion (IC) provided for a stronger and weaker response, respectively, suggesting that PAH were more readily bioavailable in PM from CC. These findings were corroborated via PAH spiking experiments indicating that IC PM contains organic components that strongly adsorb PAH thereby reducing their bioavailability. The results obtained with organic extracts in the cell transformation assay presented the highest potential for carcinogenicity in samples with high PAH contents, albeit PM from CC also demonstrated a carcinogenic potential. In conclusion, the in vitro assays employed emphasize that CC produces PM with low PAH content however with a general higher bioavailability and thus with a nearly similar carcinogenic potential than IC PM.     

Breast cancer-derived K172N,D301V mutations abolish Na/H exchanger regulatory factor 1 inhibition of platelet-derived growth factor receptor signaling

Na⁺/H⁺ exchanger regulatory factor 1 (NHERF1) is a scaffold protein known to interact with a number of cancer-related proteins. nherf1 Mutations (K172N and D301V) were recently identified in breast cancer cells. To investigate the functional properties of NHERF1, wild-type and cancer-derived nherf1 mutations were stably expressed in SKMES-1 cells respectively. NHERF1-wt overexpression suppressed the cellular malignant phenotypes, including proliferation, migration, and invasion. nherf1 Mutations (K172N and D301V) caused complete or partial loss of NHERF1 functions by affecting the PTEN/NHERF1/PDGFRβ complex formation, inactivating NHERF1 inhibition of PDGF-induced AKT and ERK activation, and attenuating the tumor-suppressor effects of NHERF1-wt. These results further demonstrated the functional consequences of breast cancer-derived nherf1 mutations (K172N and D301V), and suggested the causal role of NHERF1 in tumor development and progression.     

北虫草复合制剂对小鼠免疫功能的影响

探讨北虫草复合制剂对小鼠免疫功能的调节作用。建立小鼠免疫力降低的动物模型,实验分6组:对照组、衰老/免疫抑制模型组、白介素-2(IL-2)和北虫草复合制剂的不同剂量组。采用称重法测定免疫器官重量,计算胸腺指数和脾指数。小鼠溶血素抗体生成采用绵羊红细胞致敏法。北虫草复合制剂对小鼠脾脏指数和胸腺指数的影响:实验组(50.2±2.4与27.6±3.6)明显高于模型组(45.6±4.8与23.6±3.6),单位:(mg/10 g体重),差异有显著性(P<0.05)。对小鼠溶血素抗体生成的影响:实验组(53.53±7.8)高于药物对照组(36.50±7.3)。北虫草复合制剂能恢复衰老小鼠的胸腺指数和脾脏指数,增强免疫抑制小鼠血清溶血素含量,因此,对免疫功能低下的小鼠模型具有免疫调节作用。     

Nuclear NHERF1 expression as a prognostic marker in breast cancer

Our purpose was to investigate whether Na⁺/H⁺ exchanger regulatory factor 1 (NHERF1) expression could be linked to prognosis in invasive breast carcinomas. NHERF1, an ezrin-radixin-moesin (ERM) binding phosphoprotein 50, is involved in the linkage of integral membrane proteins to the cytoskeleton. It is therefore believed to have an important role in cell signaling associated with changes in cell cytoarchitecture. NHERF1 expression is observed in various types of cancer and is related to tumor aggressiveness. To date the most extensive analyses of the influence of NHERF1 in cancer development have been performed on breast cancer. However, the underlying mechanism and its prognostic significance are still undefined. NHERF1 expression was studied by immunohistochemistry (IHC) in a cohort of 222 breast carcinoma patients. Association of cytoplasmic and nuclear NHERF1 expression with survival was analyzed. Disease-free survival (DFS) and overall survival (OS) were determined based on the Kaplan–Meier method. Cytoplasmic NHERF1 expression was associated with negative progesterone receptor (PgR) (P=0.017) and positive HER2 expression (P=0.023). NHERF1 also showed a nuclear localization and this correlated with small tumor size (P=0.026) and positive estrogen receptor (ER) expression (P=0.010). Multivariate analysis identified large tumor size (P=0.011) and nuclear NHERF1 expression (P=0.049) to be independent prognostic variables for DFS. Moreover, the nuclear NHERF1(−)/ER(−) immunophenotype (27%) was statistically associated with large tumor size (P=0.0276), high histological grade (P=0.0411), PgR-negative tumors (P<0.0001) and high proliferative activity (P=0.0027). These patients had worse DFS compared with patients with nuclear NHERF1(+)/ER(+) tumors (75.4% versus 92.6% P=0.010). These results show that the loss of nuclear NHERF1 expression is associated with reduced survival, and the link between nuclear NHERF1 and ER expression may serve as a prognostic marker for the routine clinical management of breast cancer patients.