Rapid Mass Spectrometry Imaging to Assess the Biochemical Profile of Pituitary Tissue for Potential Intraoperative Usage.

Pituitary adenomas are relatively common intracranial neoplasms that are frequently treated with surgical resection. Rapid visualization of pituitary tissue remains a challenge as current techniques either produce little to no information on hormone-secreting function or are too slow to practically aid in intraoperative or even perioperative decision-making. Matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) represents a powerful method by which molecular maps of tissue samples can be created, yielding a two-dimensional representation of the expression patterns of small molecules and proteins from biologic samples. In this chapter, we review the use of MALDI MSI, its application to the characterization of the pituitary gland, and its potential applications for guiding the management of pituitary adenomas.

Factor VII promotes hepatocellular carcinoma progression through ERK-TSC signaling.

We previously demonstrated PAR2 starts upstreamed with tissue factor (TF) and factor VII (FVII), inhibited autophagy via mTOR signaling in HCC. However, the mechanism underlying for merging functions of PAR2 with the coagulation system in HCC progression remained unclear. The present study aimed to investigate the role of TF, FVII and PAR2 in tumor progression of HCC. The expressions of TF, FVII and PAR2 from HCC specimens were evaluated by immunohistochemical stains and western blotting. We found that the expression of FVII, but not TF and PAR2, directly related to the vascular invasion and the clinical staging. Importantly, a lower level of FVII expression was significantly associated with the longer disease-free survival. The addition of FVII but not TF induced the expression of PAR2 and phosphorylation of ERK1/2, whereas knockdown of FVII decreased PAR2 expression and ERK1/2 phosphorylation in HCC cell lines. Furthermore, levels of phosphor-TSC2 (Ser664) were increased after treatment with FVII and PAR2 agonist whereas these were significantly abolished in the presence of a potent and specific MEK/ERK inhibitor U0126. Moreover, mTOR knockdown highly reduced Hep3B migration, which could be reverted by FVII but not TF and PAR2. These results indicated that FVII/PAR2 signaling through MEK/ERK and TSC2 axis for mTOR activation has potent effects on the migration of HCC cells. In addition, FVII/PAR2 signaling elicits an mTOR-independent signaling, which promotes hepatoma cell migration in consistent with the clinical observations. Our study indicates that levels of FVII, but not TF, are associated with tumor migration and invasiveness in HCC, and provides clues that evaluation of FVII expression in HCC may be useful as a prognostic indicator in patients with HCC and may form an alternative target for further therapy.

Interconnections between autophagy and the coagulation cascade in hepatocellular carcinoma.

Autophagy has an important role in tumor biology of hepatocellular carcinoma (HCC). Recent studies demonstrated that tissue factor (TF) combined with coagulation factor VII (FVII) has a pathological role by activating a G-protein-coupled receptor called protease-activated receptor 2 (PAR2) for tumor growth. The present study aimed to investigate the interactions of autophagy and the coagulation cascade in HCC. Seventy HCC patients who underwent curative liver resection were recruited. Immunohistochemical staining and western blotting were performed to determine TF, FVII, PAR2 and light chain 3 (LC3A/B) expressions in tumors and their contiguous normal regions. We found that the levels of autophagic marker LC3A/B-II and coagulation proteins (TF, FVII and PAR2) were inversely correlated in human HCC tissues. Treatments with TF, FVII or PAR2 agonist downregulated LC3A/B-II with an increased level of mTOR in Hep3B cells; in contrast, knockdown of TF, FVII or PAR2 increased LC3A/B. Furthermore, mTOR silencing restored the impaired expression of LC3A/B-II in TF-, FVII- or PAR2-treated Hep3B cells and activated autophagy. Last, as an in vivo correlate, we administered TF, FVII or PAR2 agonist in a NOD/severe combined immunodeficiency xenograft model and showed decreased LC3A/B protein levels in HepG2 tumors with treatments. Overall, our present study demonstrated that TF, FVII and PAR2 regulated autophagy mainly via mTOR signaling. The interaction of coagulation and autophagic pathways may provide potential targets for further therapeutic application in HCC.

Regulation of heme oxygenase 1 expression by miR-27b with stem cell therapy for liver regeneration in rats.

Adipose-derived mesenchymal stem cells (ASCs) have been considered to be attractive and readily available adult mesenchymal stem cells (MSCs) and are becoming increasingly popular for use in regenerating cell therapy. However, recent evidence attributed a fibrotic potential to MSCs which differentiated into myofibroblasts with highly increased α-smooth muscle actin (α-SMA) expression while transplanted into an injured/regenerating liver in mice. In this study, we studied the role of miR-27b in ASCs and their regenerative potential after partial liver resection in rats. ASCs transfected with control siRNA or miR-27b were intravenously injected into autologous rats undergoing 70% partial hepatectomy (PH). Our data showed that the regenerative capacities of ASCs with overexpressed miR-27b were significantly higher compared with control ASCs. However, the enhanced regeneration, hepatic differentiation, and suppressed liver inflammation, as well as fibrotic activity, were significantly reverted by ZnPP coadministration (heme oxygenase-1 [HO-1] inhibitor) indicating an important role of HO-1 in the regenerating and cytoprotective activities of miR-27b-transfected ASCs. We demonstrated that administration of autologous ASCs overexpressed with miR-27b enhances rapid and early liver regeneration and, importantly, preserves function after PH. The ASCs with miR-27b overexpression might offer a viable therapeutic option to facilitate rapid recovery after liver resection.

miR-499A>G rs3746444 and miR-146aG>C expression and hepatocellular carcinoma risk in the Chinese population.

We conducted a case-control study of a possible association of miR-499A>G rs3746444 and miR-146aG>C rs2910164 with risk of hepatocellular carcinoma. Samples from 172 hepatocellular carcinoma patients and 185 cancer-free controls were collected from October 2008 to December 2011. PCR-RFLP analysis was performed to determine the polymorphisms in each individual. The MAFs of miR-146aG>C and miR-499A>G in controls were similar to that known from the SNP database, and frequencies of genotypes in controls were in line with Hardy-Weinberg equilibrium. We found that miR-499 AG was significantly associated with decreased risk for hepatocellular carcinoma when compared with miR-499 AA genotype (adjusted odds ration = 0.74, 95% confidence interval = 0.24-0.96). However, subjects carrying miR-146a GG had a non-significant 0.62-fold decreased risk of hepatocellular carcinoma. We did not find a significant association of miR-146aG>C rs2910164 and miR-499A>G rs3746444 polymorphisms with hepatocellular carcinoma risk in the Chinese population. Further investigations are warranted to clarify the relationship between miRNA polymorphisms and susceptibility to hepatocellular carcinoma risk in various ethnic populations.

Evidence for inhibition of low density lipoprotein oxidation and cholesterol accumulation by apolipoprotein H (beta2-glycoprotein I).

Low density lipoprotein (LDL) oxidation and lipid accumulation are thought to enhance the progression of atherosclerosis. Apolipoprotein H (apoH) has been implicated in the development of human atherosclerosis. However, the roles of apoH in the oxidative modification of LDL and cellular accumulation of lipid constituents remained uncharacterized. In this study, the level of plasma apoH was found to be significantly associated with the oxidative susceptibility of LDL in human subjects. Plasma levels of apoH were positively correlated with the lag time but negatively correlated with LDL oxidation rate in conjugated diene formation. By using a J774 A.1 macrophage culture system, we found that apoH could not only inhibit the formation of conjugated diene and thiobarbituric acid-reactive substances, but also reduce the electrophoretic mobility of oxidized LDL. Furthermore, apoH decreased cellular accumulation of cholesterol via a reduction in cholesterol influx and an increase in cholesterol efflux. This is the first demonstration that apoH appears to have "antioxidant"-like effects on LDL oxidation. The results also suggest that apoH can inhibit the translocation of cholesterol from extracellular pools to macrophages, suggesting that apoH may play an important role in the prevention of atherosclerosis.

Clinical application of intratumoral blood flow study in patients with endometrial carcinoma.

BACKGROUND: The objective of this study was to evaluate the correlation between intratumoral blood flow as assessed by color Doppler ultrasound with stage, tumor grade, depth of invasion, and lymph node metastasis in endometrial carcinoma and determine its clinical usefulness. METHODS: Sixty-six patients with endometrial carcinoma were enrolled. All patients received surgical treatment. Transvaginal color Doppler ultrasound was performed before surgery to detect the arterial blood flow signals within the tumors and the lowest resistance index (RI) was recorded. Formalin fixed, paraffin embedded pathology slides were reviewed by a senior pathologist to evaluate the histologic grading, tumor size, depth of myometrial invasion, and presence of lymph node metastasis. RESULTS: Intratumoral RI correlated well with surgical staging, histologic grading, the depth of myometrial invasion, and the presence of lymph node metastasis. Significantly lower RI was noted in tumors of advanced stage (> than International Federation of Obstetrics and Gynecology [FIGO] Stage I) (0.38 +/- 0.09 vs. 0.54 +/- 0.11; P < 0.001), tumors with higher histologic grade (Grade 3) (0.36 +/- 0.08 vs. 0.53 +/- 0.11; P < 0.001), tumors with deep myometrial invasion (> 50% myometrial thickness) (0.38 +/- 0.07 vs. 0.54 +/- 0.11; P < 0.001), and tumors with lymph node metastasis (0.34 +/- 0.07 vs. 0.52 +/- 0.11; P < 0.001) compared with tumors with Stage I, Grade 1/2 histology, no or superficial myometrial invasion, and absence of lymph node metastasis, respectively. CONCLUSIONS: Intratumoral blood flow analysis assessed by color Doppler ultrasound correlates well with surgical stage, tumor grade, myometrial invasion, and lymph node metastasis in patients with endometrial carcinoma. Assessment of tumor angiogenesis using color Doppler ultrasound provides useful information for the preoperative prediction regarding stage, histologic grade, depth of myometrial invasion, and presence of lymph node metastasis in patients with endometrial carcinoma. The authors believe routine pelvic lymph node dissection should be performed for those patients whose lowest RI values of intratumoral blood flow are < or = 0.4 because those patients are at very high risk for pelvic lymph node involvement.

Preoperative ultrasound study in predicting lymph node metastasis for endometrial cancer patients.

OBJECTIVE: The objective of this study was to evaluate the efficacy of preoperative ultrasound (US) findings such as tumor size, status of myometrial invasion, and intratumoral "resistance index" (RI) in predicting lymph node metastasis in endometrial carcinoma patients. METHODS: Forty-two patients with endometrial cancer were enrolled. All patients underwent total abdominal hysterectomy, pelvic lymph node dissection or sampling, and para-aortic lymph node sampling. Two-dimensional and color Doppler US were performed before surgery to measure tumor size, depth of myometrial invasion, and intratumoral arterial RI. Formalin-fixed, paraffin-embedded pathologic slides from surgical specimens were reviewed by a senior pathologist to evaluate histologic type and grade, depth of myometrial invasion, cervical involvement, lymph-vascular emboli, and status of lymph node metastasis. RESULTS: There were 12 patients with pelvic and/or para-aortic lymph node metastases and 30 patients without nodal metastases. Patients with tumors larger than 2.5 cm by US (11/12 vs 14/30, P = 0.008), more than half myometrial invasion by US (9/12 vs 5/30, P < 0.001), and intratumoral RI values less than 0.4 by US (12/12 vs 4/30, P < 0.001) had a significantly higher incidence of nodal metastases as compared with patients with tumors smaller than 2.5 cm, no or superficial myometrial invasion, and RI values higher than 0.4, respectively. Multiple regression analysis showed that only intratumoral RI values less than 0.4 were significantly correlated with nodal metastasis (P < 0.001, r2 = 0. 650). We used the intratumoral RI value as the parameter to evaluate nodal metastasis in endometrial cancer patients. Twelve of sixteen patients with intratumoral RI values <0.4 had a high incidence of nodal metastases. None of the 26 patients with intratumoral RI values >0.4 had nodal metastases. CONCLUSIONS: Preoperative ultrasound features can offer important information for predicting lymph node metastasis in endometrial cancer patients. Patients with tumors with intratumoral RI values less than 0.4 should be highly suspected of having lymph node metastases and further management such as pelvic lymph node dissection or postoperative pelvic radiotherapy would be needed for these patients.

Endometrial stromal sarcoma of twenty cases.

BACKGROUND: Endometrial stromal sarcoma is a rare neoplasm. We reviewed twenty cases to study the characteristics of this disease. METHODS: Twenty cases of endometrial stromal sarcoma were treated at our hospital. The clinical stage, treatment and outcome were retrospectively analyzed. RESULTS: Endometrial stromal sarcoma comprised 4.3% of corpus cancers and 46.4% of uterine sarcomas at our hospital. Seven cases were stage I, one was stage II, ten were stage III, and two were stage IV at the time of diagnosis. Histopathologically, seventeen cases were classified as low-grade sarcoma and three were high-grade sarcoma. Seven patients had recurrence and five of them had already died of disease. Among these recurrent patients, one was stage II and six were stage III. All three patients with high-grade sarcoma and four with low-grade sarcoma had recurrence. CONCLUSIONS: We think mitotic count is an important prognostic factor in low-grade endometrial stromal sarcoma and high-grade endometrial stromal sarcoma has a poor prognosis even with post-operative adjuvant treatment.

Sonographic characteristics of adenofibroma of the endometrium following tamoxifen therapy for breast cancer: two case reports.

Adenofibroma of the endometrium is thought to be a rare benign variant of the mixed mesodermal tumor, and its preoperative diagnosis is difficult. We describe the sonographic characteristics of two cases of adenofibroma of the endometrium. In both cases the patient was receiving prolonged tamoxifen therapy following surgery for breast cancer. Sonographically, this rare disease is observed as an intracavitary mass containing multiple small cysts with low-resistance intratumor blood flow. The unique sonographic findings make the preoperative diagnosis possible.

Functional association of essential splicing factor(s) with PRP19 in a protein complex.

We have previously shown that the yeast PRP19 protein is a spliceosomal component, but is not tightly associated with small nuclear RNAs. It appears to associate with the spliceosome concomitant with or just after dissociation of the U4 small nuclear RNA during spliceosome assembly. We have found that PRP19 is associated with a protein complex in the splicing extract and that at least one of the associated components is essential for splicing. Taking advantage of the epitope tagging technique, we have isolated the PRP19-associated complex by affinity chromatography. The isolated complex is functional for complementation for the heat-inactivated prp19 mutant extract, and consists of at least seven polypeptides in addition to PRP19. At least three of these can interact directly with the PRP19 protein. We also show that the PRP19 protein itself is in an oligomeric form, which might be a prerequisite for its interaction with these proteins.