Aquaporin-1 and -5 are involved in the invasion and proliferation of soft tissue sarcomas

Background and aim

Recent studies of several carcinomas have reported that aquaporin possesses novel oncogenic properties. The aim of this study was to clarify the involvement of aquaporin-1 and ?5 in the proliferation, invasion and metastasis of soft tissue sarcomas.

Clinical and Immunological Characterization of ICF Syndrome in Japan

Objective

Immunodeficiency, centromeric instability, and facial anomalies (ICF) syndrome is a rare autosomal recessive primary immunodeficiency. Hypogammaglobulinemia is a major manifestation of ICF syndrome, but immunoglobulin replacement therapy does not seem to be effective for some ICF patients. Therefore, we aimed to reassess the immunological characteristics of this syndrome.

Methods

Eleven Japanese patients with ICF syndrome were enrolled. We performed whole-exome sequencing in four cases and homozygosity mapping using SNP analysis in two. We evaluated their clinical manifestations and immunological status.

Results

We newly diagnosed six ICF patients who had tentatively been diagnosed with common variable immunodeficiency. We identified two novel mutations in the DNMT3B gene and one novel mutation in the ZBTB24 gene. All patients showed low serum IgG and/or IgG₂ levels and were treated by periodic immunoglobulin replacement therapy. Three of the six patients showed worse results of the mitogen-induced lymphocyte proliferation test. Analyses of lymphocyte subpopulations revealed that CD19⁺CD27⁺ memory B cells were low in seven of nine patients, CD3⁺ T cells were low in three patients, CD4/8 ratio was inverted in five patients, CD31⁺ recent thymic emigrant cells were low in two patients, and CD19⁺ B cells were low in four patients compared with those in the normal controls. ICF2 patients showed lower proportions of CD19⁺ B cells and CD16⁺56⁺ NK cells and significantly higher proportions of CD3⁺ T cells than ICF1 patients. T cell receptor excision circles were undetectable in two patients. Despite being treated by immunoglobulin replacement therapy, three patients died of influenza virus, fatal viral infection with persistent Epstein–Barr virus infection, or JC virus infection. One of three dead patients showed normal intelligence with mild facial anomaly. Two patients presented with autoimmune or inflammatory manifestations. Infectious episodes decreased in three patients who were started on trimethoprim–sulfamethoxazole and/or antifungal drugs in addition to immunoglobulin replacement therapy. These patients might have suffered from T cell immunodeficiency.

Conclusion

These results indicate that patients with ICF syndrome have a phenotype of combined immunodeficiency. Thus, to achieve a better prognosis, these patients should be treated as having combined immunodeficiency in addition to receiving immunoglobulin replacement therapy.

     

Whey protein intake after resistance exercise activates mTOR signaling in a dose-dependent manner in human skeletal muscle

Purpose

Protein ingestion after resistance exercise increases muscle protein synthesis (MPS) in a dose-dependent manner. However, the molecular mechanism(s) for the dose-dependency of MPS remains unclear. This study aimed to determine the dose response of mammalian target of rapamycin (mTOR) signaling in muscle with ingestion of protein after resistance exercise.

Methods

Fifteen male subjects performed four sets of six unilateral isokinetic concentric knee extensions. Immediately after exercise, eight subjects consumed water only. The other seven subjects, in a randomized-order crossover design, took either a 10 [3.6 g essential amino acids (EAA)] or 20 g (7.1 g EAA) solution of whey protein. Muscle biopsies from the vastus lateralis muscle were taken 30 min before and 1 h after resistance exercise. Phosphorylation of Akt (Ser473), mTOR (Ser2448), 4E-BP1 (Thr37/46), and S6K1 (Thr389) was measured by western blotting.

Results

Concentric knee extension exercise alone did not increase phosphorylation of Akt and mTOR 1 h after exercise, but ingesting protein after exercise significantly increased the phosphorylation of Akt and mTOR in a dose-dependent manner (P < 0.05). 4E-BP1 phosphorylation significantly decreased after resistance exercise (P < 0.05), but subjects who took 10 or 20 g of protein after exercise showed increased 4E-BP1 from post-exercise dephosphorylation (P < 0.05). S6K1 phosphorylation significantly increased after resistance exercise (P < 0.05), and 20 g of protein further increased S6K1 phosphorylation compared with ingestion of 10 g (P < 0.05).

Conclusions

These findings suggest that whey protein intake after resistance exercise activates mTOR signaling in a dose-dependent manner in untrained men.     

Features and prognostic impact of distant metastasis in patients with stage IV lung adenocarcinoma harboring EGFR mutations: importance of bone metastasis

Mutated epidermal growth factor receptor (EGFR) and signaling pathways were associated with multiple brain and intra-pulmonary metastases, oncogenic progression and metastasis. However, features of metastasis to other organs and the independent prognostic influence of metastatic lesions were not elucidated in patients with lung cancer harboring EGFR mutations. Between January 2007 and April 2012, we treated 277 patients diagnosed with stage IV lung adenocarcinoma. Studied were 246 patients with available tumor EGFR mutation data who also underwent radiographic evaluation of lung, abdominal, brain, and bone metastases. The EGFR mutated group (N = 98) had significantly more metastatic lesions in the brain and bone than the wild-type group (N = 148): brain, 3 (1–93) versus 2 (1–32) median (range), P = 0.023; bone, 3 (1–43) versus 2 (1–27), P = 0.035, respectively. In addition, EGFR mutations were significantly more frequent in patients with multiple than non-multiple lung metastases (24/40 vs. 12/42, P = 0.004). Multivariate analysis showed that bone metastasis was a significant independent negative predictive factor of overall survival (OS) in patients with mutated [hazard ratio (HR) 2.04; 95 % confidence interval (CI) 1.17–3.64; P = 0.011] and wild-type EGFR (HR 2.09; 95 % CI 1.37–3.20; P < 0.001). In conclusion, patients with mutated EGFR had more lung, brain, and bone metastases, and bone metastasis was an independent negative predictor of OS.     

Enhanced <Emphasis Type="Italic">AZIN1</Emphasis> RNA editing and overexpression of its regulatory enzyme ADAR1 are important prognostic biomarkers in gastric cancer

Background

Adenosine-to-inosine (A-to-I) RNA editing is catalyzed by adenosine deaminases acting on RNA (ADAR) enzymes. Recent evidence suggests that RNA editing of antizyme inhibitor 1 (AZIN1) RNA is emerging as a key epigenetic alteration underlying cancer pathogenesis.

Methods

We evaluated AZIN1 RNA editing levels, and the expression of its regulator, ADAR1, in 280 gastric tissues from 140 patients, using a RNA editing site-specific quantitative polymerase chain reaction assays. We also analyzed the clinical significance of these results as disease biomarkers in gastric cancer (GC) patients.

Results

Both AZIN1 RNA editing levels and ADAR1 expression were significantly elevated in GC tissues compared with matched normal mucosa (P?<?0.0001, 0.0008, respectively); and AZIN1 RNA editing was positively correlated with ADAR1 expression. Elevated expression of ADAR1 significantly correlated with poor overall survival (P?=?0.034), while hyper-edited AZIN1 emerged as an independent prognostic factor for OS and disease-free survival in GC patients [odds ratio (OR):1.98, 95% CI 1.17–3.35, P?=?0.011, OR: 4.55, 95% CI 2.12–9.78, P?=?0.0001, respectively]. Increased AZIN1 RNA editing and ADAR1 over-expression were significantly correlated with key clinicopathological factors, such as advanced T stage, presence of lymph node metastasis, distant metastasis, and higher TNM stages in GC patients. Logistic regression analysis revealed that hyper-editing status of AZIN1 RNA was an independent risk factor for lymph node metastasis in GC patients [hazard ratio (HR):3.03, 95% CI 1.19–7.71, P?=?0.02]. Conclusions: AZIN1 RNA editing levels may be an important prognostic biomarker in GC patients, and may serve as a key clinical decision-making tool for determining preoperative treatment strategies in GC patients.

     

Polymeric photosensitizer-embedded self-expanding metal stent for repeatable endoscopic photodynamic therapy of cholangiocarcinoma

Photodynamic therapy (PDT) is a new therapeutic approach for the palliative treatment of malignant bile duct obstruction. In this study, we designed photosensitizer-embedded self-expanding nonvascular metal stent (PDT-stent) which allows repeatable photodynamic treatment of cholangiocarcinoma without systemic injection of photosensitizer. Polymeric photosensitizer (pullulan acetate-conjugated pheophorbide A; PPA) was incorporated in self-expanding nonvascular metal stent. Residence of PPA in the stent was estimated in buffer solution and subcutaneous implantation on mouse. Photodynamic activity of PDT-stent was evaluated through laserexposure on stent-layered tumor cell lines, HCT-116 tumor-xenograft mouse models and endoscopic intervention of PDT-stent on bile duct of mini pigs. Photo-fluorescence imaging of the PDT-stent demonstrated homogeneous embedding of polymeric Pheo-A (PPA) on stent membrane. PDT-stent sustained its photodynamic activities at least for 2 month. And which implies repeatable endoscopic PDT is possible after stent emplacement. The PDT-stent after light exposure successfully generated cytotoxic singlet oxygen in the surrounding tissues, inducing apoptotic degradation of tumor cells and regression of xenograft tumors on mouse models. Endoscopic biliary in-stent photodynamic treatments on minipigs also suggested the potential efficacy of PDT-stent on cholangiocarcinoma. In vivo and in vitro studies revealed our PDT-stent, allows repeatable endoscopic biliary PDT, has the potential for the combination therapy (stent plus PDT) of cholangiocarcinoma.     

Muscle‐specific calpain‐3 is phosphorylated in its unique insertion region for enrichment in a myofibril fraction

CAPN3 (also called p94/calpain‐3) is a skeletal muscle‐specific calpain, an intracellular cysteine protease. Loss of CAPN3 protease activity and/or structural functions cause limb‐girdle muscular dystrophy type 2A (LGMD2A). However, the precise mechanism of action of CAPN3 in skeletal muscles in vivo remains largely elusive. By studying the protein modifications that regulate CAPN3 activity, we found that CAPN3 was phosphorylated. By performing mutagenesis and mass spectrometry analyses, we identified two Ser residues at positions 629 and 636 in human CAPN3 that are phosphorylated and showed that S629 is a major phosphorylation site. Intriguingly, rapid and exhaustive autolysis of CAPN3 was slightly attenuated by the substitution of S629. In skeletal muscles, phosphorylated CAPN3 was enriched in the myofibril fraction. These results imply that phosphorylated CAPN3 is a myofibril structural component and/or participates in myofibril‐based signaling pathways, rather than functions as a protease. We evaluated the relationship between phosphorylated CAPN3 and the pathology of LGMD2A. The level of phosphorylated CAPN3 was greatly reduced in LGMD2A muscles. Our findings suggest that phosphorylated CAPN3 is involved in the pathology of LGMD2A through defects in myofibril integrity and/or signaling pathways. This is the first report that phosphorylation of CAPN3 may be involved in its physiological function.     

Adenocarcinoma arising in small sessile serrated adenoma/polyp (SSA/P) of the colon: Clinicopathological study of eight lesions

We reviewed the clinicopathological findings of eight cases of sessile serrated adenoma/polyps (SSA/Ps) with carcinoma, the largest diameter of which was 10 mm or less. All lesions were polyps located in the right side of the colon. Four lesions showed submucosal invasion and one lesion invaded the proper muscle layer. The depth of invasion, however, did not seem to be related to the carcinoma area size. Most carcinomas were well to moderately differentiated tubular adenocarcinomas focally showing some serrated appearances, and the predominant component of one carcinoma was a poorly differentiated medullary growth with inflammatory stroma. Rapid progression to invasive carcinoma from SSA/P was suggested for the carcinoma with proper muscle invasion whereas one submucosally invasive carcinoma was considered to progress over 7 years. Immunohistochemically, it was suggested that with or without hMLH1 protein loss, alterations of p53 and/or Wnt signaling pathway can be involved in the cancerization through SSA/Ps. The carcinomas irregularly imitated the mucin expression of the SSA/Ps (positive for MUC5AC and MUC2, and MUC6 expression in crypt bases), which was lost with progression of the carcinomas. Analyses of small SSA/P lesions with cancerization would facilitate the understanding of the mode of progression of SSA/Ps and their early detection.     

Microfibril-Associated Glycoprotein-1 Controls Human Ciliary Zonule Development In Vitro

The ciliary zonule in the eye, also known as Zinn’s zonule, is composed of oxytalan fibers, which are bundles of microfibrils consisting mainly of fibrillin-1. However, it is still unclear which of the microfibril-associated molecules present in the ciliary zonule controls oxytalan fibers. Microfibril-associated glycoprotein-1 (MAGP-1) is the only microfibril-associated molecule identified in the human ciliary zonule. In the present study, we used siRNA against MAGP-1 in cultures of human non-pigmented ciliary epithelial cells to examine the extracellular deposition and appearance of fibrillin-1 employing Western blotting and immunofluorescence. MAGP-1 suppression led to a reduction of fibrillin-1 deposition. Immunofluorescence also confirmed that RNAi-mediated down-regulation of MAGP-1 led to suppression of fiber development. These results suggest that MAGP-1 plays a crucial role in the extracellular deposition of fibrillin-1 during formation of the human ciliary zonule.