Faculty Bibliography

Epigallocatechin gallate (EGCG) of green tea and the nutraceutical CystiCran^®-40 (containing 40% proanthocyanidins) of the cranberry plant have been associated with antiviral activity. The purpose of this work was to determine the mechanism of antiviral synergy between each compound. Coliphage T4II (phage T4) and the rotavirus strain SA-11(RTV) were used as model virus systems. Individual and combined flavonoids structural and molecular weight analyses were performed by NMR and HPCL/MS, respectively. A suboptimal concentration of EGCG or C-40 alone or in combination reduced phage infectivity by ≤10%. Similarly, EGCG (30 µg/ml) and C-40 (25 µg/ml), respectively, reduced RTV titers by 3 and 13%. However, RTV titers were reduced by 32% (p < .05) with both flavonoids used in combination. RTV was not recognized in host cells by electron microscopy 24-h post-inoculation. NMR and HPLC/MS findings revealed significant structural and potential changes in molecular weight of the flavonoids in complex.

BACKGROUND & AIMS: Obesity-induced non-alcoholic fatty liver disease (NAFLD) develops, in part, via excess insulin-stimulated hepatic de novo lipogenesis, which increases, paradoxically, in patients with obesity-induced insulin resistance. Pleckstrin homology domain leucine-rich repeat protein phosphatase 2 (PHLPP2) terminates insulin signaling by dephosphorylating Akt; levels of PHLPP2 are reduced in livers from obese mice. We investigated whether loss of hepatic PHLPP2 is sufficient to induce fatty liver in mice, mechanisms of PHLPP2 degradation in fatty liver, and expression of genes that regulate PHLPP2 in livers of patients with NAFLD. METHODS: C57BL/6J mice (controls), obese db/db mice and mice with liver-specific deletion of PHLPP2 (L-PHLPP2) fed either normal chow or high-fat diet (HFD) were analyzed for metabolic phenotypes including glucose tolerance and hepatic steatosis. PHLPP2-deficient primary hepatocytes or CRISPR/Cas9-mediated PHLPP2-knockout hepatoma cells were analyzed for insulin signaling and gene expression. We performed mass spectrometry analyses of livers tissues from C57BL/6J mice transduced with Ad-HA-FLAG-PHLPP2 to identify post-translational modifications to PHLPP2 and proteins that interact with PHLPP2. We measured levels of mRNAs by quantitative reverse transcription PCR in liver biopsies from patients with varying degrees of hepatic steatosis. RESULTS: PHLPP2-knockout hepatoma cells and hepatocytes from L-PHLPP2 mice showed normal initiation of insulin signaling, but prolonged insulin action. Chow-fed L-PHLPP2 mice had normal glucose tolerance but hepatic steatosis. In HFD-fed C57BL/6J or db/db obese mice, endogenous PHLPP2 was degraded by glucagon and PKA-dependent phosphorylation of PHLPP2 (at Ser1119 and Ser1210), which led to PHLPP2 binding to potassium channel tetramerization domain containing 17 (KCTD17), a substrate-adaptor for Cul3-RING ubiquitin ligases. Levels of KCTD17 mRNA were increased in livers of HFD-fed C57BL/6J or db/db obese mice and in liver biopsies patients with NAFLD, compared with liver tissues from healthy control mice or patients without steatosis. Knockdown of KCTD17 with small hairpin RNA in primary hepatocytes increased PHLPP2 protein but not Phlpp2 mRNA, indicating that KCTD17 mediates PHLPP2 degradation. KCTD17 knockdown in obese mice prevented PHLPP2 degradation and decreased expression of lipogenic genes. CONCLUSIONS: In mouse models of obesity, we found that PHLPP2 degradation induced lipogenesis without affecting gluconeogenesis. KCTD17, which is upregulated in liver tissues of obese mice and patients with NAFLD, binds to phosphorylated PHLPP2 to target it for ubiquitin-mediated degradation; this increases expression of genes that regulate lipogenesis to promote hepatic steatosis. Inhibitors of this pathway might be developed for treatment of patients with NAFLD.

Coloboma of the eye, Heart defects, Atresia of the choanae, Retardation of growth and/or development, Genital and/or urinary abnormalities, and Ear abnormalities and deafness (CHARGE) syndrome is a disorder with multiple congenital anomalies seen on imaging. A retrospective review of 10 patients with CHARGE syndrome who underwent MR imaging of the brain as part of a preoperative evaluation for cochlear implantation was conducted. Structural abnormalities of the entire MR imaging of the head were evaluated, including the auditory system, olfactory system, face, skull base, and central nervous system. The most frequent MR imaging findings included dysplasias of the semicircular canals and hypoplasia of the frontal lobe olfactory sulci. Less frequent findings included cleft lip/palate and coloboma. Our study uncovered new findings of a J-shaped sella, dorsal angulation of the clivus, and absent/atrophic parotid glands, not previously described in patients with CHARGE. Our results emphasize the utility of MR imaging in the diagnosis and management of patients with CHARGE syndrome.

Due to their anatomical location, vocal folds are highly susceptible to injury from external and internal stressors that can lead to irreversible damage and changes in function. As the structure and composition of the vocal folds are heavily linked to their unique function, we hypothesize that a vocal fold-derived extracellular matrix (ECM) would be the ideal scaffold in a regenerative approach to vocal fold repair. Our group has previously described a porcine-derived vocal fold lamina propria ECM (VFLP-ECM)¹. In order to optimize the delivery modality of the VFLP-ECM, we have developed an injectable hydrogel form of the ECM scaffold and have studied the effects of tissue specificity using human vocal fold fibroblasts (hVFF) and human peripheral blood-derived macrophages (hPB-Macrophages). Both cell types play unique roles during the inflammatory and wound healing response at the site of vocal fold injury. In the present study, we compare VFLP-ECM with other ECM hydrogels (such as collagen, heart, bladder) in their ability to activate and modify gene expression of hVFFs and hPB-macrophages. This information will help us tailor the VFLP-ECM hydrogel to mod-ulate the environment present during vocal fold injury