Aquaporins in Cancer Biology.

Aquaporins (AQPs) are a family of transmembrane water channel proteins, which were initially characterized as a novel protein family that plays a vital role in transcellular and transepithelial water movement. AQP1, AQP2, AQP4, AQP5, and AQP8 are primarily water selective, whereas AQP3, AQP7, AQP9, and AQP10 (called "aqua-glyceroporins") also transport glycerol and other small solutes. Recently, multiple reports have suggested that AQPs have important roles in cancer cell growth, migration, invasion, and angiogenesis, each of which is important in human carcinogenesis. Here, we review recent data concerning the involvement of AQPs in tumor growth, angiogenesis, and metastasis and explore the expression profiles from various resected cancer samples to further dissect the underlying molecular mechanisms. Moreover, we discuss the potential role of AQPs during the development of genomic instability and performed modeling to describe the integration of binding between AQPs with various SH3 domain binning adaptor molecules. Throughout review and discussion of numerous reports, we have tried to provide key evidence that AQPs play key roles in tumor biology, which may provide a unique opportunity in designing a novel class of anti-tumor agents.

Induction of GD3/α1-adrenergic receptor/transglutaminase 2-mediated erythroid differentiation in chronic myelogenous leukemic K562 cells.

The disialic acid-containing glycosphingolipid GD3 recruited membrane transglutaminase 2 (TG2) as a signaling molecule for erythroid differentiation in human chronic myelogenous leukemia (CML) K562 cells. The α1-adrenergic receptor (α1-AR)/TG2-mediated signaling pathway regulated GD3 functions, including gene expression and production, to differentiate CML K562 cells into erythroid lineage cells. Epinephrine, an AR agonist, increased membrane recruitment as well as GTP-photoaffinity of TG2, inducing GD3 synthase gene expression. Epinephrine activated PI3K/Akt signaling and GTPase downstream of TG2 activated Akt. The coupling of TG2 and GD3 production was specifically suppressed by prazosin (α1-AR antagonist), but not by propranolol (ß-AR antagonist) or rauwolscine (α2-AR antagonist), indicating α1-AR specificity. Small interfering RNA (siRNA) experiment results indicated that the α1-AR/TG2-mediated signaling pathway activated PKCs α and δ to induce GD3 synthase gene expression. Transcription factors CREB, AP-1, and NF-κB regulated GD3 synthase gene expression during α1-AR-induced differentiation in CML K562 cells. In addition, GD3 synthase gene expression was upregulated in TG2-transfected cells via α1-AR with expression of erythroid lineage markers and benzidine-positive staining. α1-AR/TG2 signaling pathway-directed GD3 production is a crucial step in erythroid differentiation of K562 cells and GD3 interacts with α1-AR/TG2, inducing GD3/α1-AR/TG2-mediated erythroid differentiation. These results suggest that GD3, which acts as a membrane mediator of erythroid differentiation in CML cells, provides a therapeutic avenue for leukemia treatment.

Monosialyl Ganglioside GM3 Decreases Apolipoprotein B-100 Secretion in Liver Cells.

Some sialic acid-containing glycolipids are known to regulate development of atherosclerosis with accumulated plasma apolipoprotein B-100 (Apo-B)-containing lipoproteins, because Apo-B as an atherogenic apolipoprotein is assembled mainly in VLDL and LDL. Previously, we have elucidated that disialyl GD3 promotes the microsomal triglyceride transfer protein (MTP) gene expression and secretion of triglyceride (TG)-assembled ApoB, claiming the GD3 role in ApoB lipoprotein secretion in liver cells. In the synthetic pathway of gangliosides, GD3 is synthesized by addition of a sialic acid residue to GM3. Thus, there should be some regulatory links between GM3 and GD3. In this study, exogenous and endogenous monosialyl GM3 has been examined how GM3 plays a role in ApoB secretion in Chang liver cells in a view point of MTP and ApoB degradation in the same cells. The level of GM3 ganglioside in the GM3 synthase gene-transfected cells was increased in the cell extract, but not in the medium. In addition, GM3 synthase gene-transfected cells showed a diminished secretion of TG-enriched ApoB with a lower content of TG in the medium. Exogenous GM3 treatment for 24 h exerted a dose dependent inhibitory effect on ApoB secretion together with TG, while a liver-specific albumin was unchanged, indicating that GM3 effect is limited to ApoB secretion. GM3 decreased the mRNA level of MTP gene, too. ApoB protein assembly dysregulated by GM3 indicates the impaired ApoB secretion is caused by a proteasome-dependent pathway. Treatment with small interfering RNAs (siRNAs) decreased ApoB secretion, but GM3-specific antibody did not. These results indicate that plasma membrane associated GM3 inhibits ApoB secretion, lowers development of atherosclerosis by decreasing the secretion of TG-enriched ApoB containing lipoproteins, suggesting that GM3 is an inhibitor of ApoB and TG secretion in liver cells. J. Cell. Biochem. 118: 2168-2181, 2017. © 2017 Wiley Periodicals, Inc.

Risk of malignancy in thyroid incidentalomas identified by fluorodeoxyglucose-positron emission tomography.

BACKGROUND: Thyroid incidentalomas detected by 2-deoxy-2-18F-fluoro-D-glucose positron emission tomography/computed tomography (18F-FDG PET/CT) have been reported in 1% to 4% of the population, with a risk of malignancy of 27.8% to 74%. We performed a retrospective review of FDG-avid thyroid incidentalomas in cancer screening subjects and patients with nonthyroid cancer. The risk of malignancy in thyroid incidentaloma and its association with the maximal standardized uptake value (SUVmax) in 18F-FDG PET/CT were evaluated to define the predictor variables in assessing risk of malignancy. METHODS: A total of 2,584 subjects underwent 18F-FDG PET/CT for metastatic evaluation or cancer screening from January 2005 to January 2010. Among them, 36 subjects with FDG-avid thyroid incidentalomas underwent further diagnostic evaluation (thyroid ultrasonography-guided fine needle aspiration cytology [FNAC] or surgical resection). We retrospectively reviewed the database of these subjects. RESULTS: Of the 2,584 subjects who underwent 18F-FDG PET/CT (319 for cancer screening and 2,265 for metastatic evaluation), 52 (2.0%) were identified as having FDG-avid thyroid incidentaloma and cytologic diagnosis was obtained by FNAC in 36 subjects. Of the subjects, 15 were proven to have malignant disease: 13 by FNAC and two by surgical resection. The positive predictive value of malignancy in FDG-avid thyroid incidentaloma was 41.7%. Median SUVmax was higher in malignancy than in benign lesions (4.7 [interquartile range (IQR), 3.4 to 6.0] vs. 2.8 [IQR, 2.6 to 4.0], P=0.001). CONCLUSION: Thyroid incidentalomas found on 18F-FDG PET/CT have a high risk of malignancy, with a positive predictive value of 41.7%. FDG-avid thyroid incidentalomas with higher SUVmax tended to be malignant.

Metastasis of colon cancer to medullary thyroid carcinoma: a case report.

Metastasis to the primary thyroid carcinoma is extremely rare. We report here a case of colonic adenocarcinoma metastasis to medullary thyroid carcinoma in a 53-yr old man with a history of colon cancer. He showed a nodular lesion, suggesting malignancy in the thyroid gland, in a follow-up examination after colon cancer surgery. Fine needle aspiration biopsy (FNAB) of the thyroid gland showed tumor cell clusters, which was suspected to be medullary thyroid carcinoma (MTC). The patient underwent a total thyroidectomy. Using several specific immunohistochemical stains, the patient was diagnosed with colonic adenocarcinoma metastasis to MTC. To the best of our knowledge, the present patient is the first case of colonic adenocarcinoma metastasizing to MTC. Although tumor-tumor metastasis to primary thyroid carcinoma is very rare, we still should consider metastasis to the thyroid gland, when a patient with a history of other malignancy presents with a new thyroid finding.

Clinical Characteristics and Metabolic Features of Patients with Adrenal Incidentalomas with or without Subclinical Cushing's Syndrome.

BACKGROUND: The aim of this study was to examine the clinical characteristics of adrenal incidentalomas discovered by computed tomography (CT) and to investigate metabolic features of subclinical Cushing's syndrome (SCS) in patients with adrenal incidentalomas in a tertiary hospital in Korea. METHODS: This retrospective study examined the clinical aspects of 268 patients with adrenal incidentalomas discovered by CT at Soonchunhyang University Bucheon Hospital. Clinical data and endocrine function of the patients as well as histological findings were obtained from medical records, while anatomic characteristics were analyzed by reviewing imaging studies. Hormonal tests for pheochromocytoma, Cushing's syndrome, and aldosterone-secreting adenoma were performed. RESULTS: Most (n=218, 81.3%) cases were nonfunctioning tumors. Of the 50 patients with functioning tumors (18.7%), 19 (7.1%) were diagnosed with SCS, nine (3.4%) with overt Cushing's syndrome, 12 (4.5%) with primary aldosteronism, and 10 (3.7%) with pheochromocytoma. Malignant tumors (both primary and metastatic) were rare (n=2, 0.7%). Body mass index, fasting glucose, hemoglobin A1c, and total cholesterol were significantly higher in patients with SCS in comparison with those with nonfunctioning tumors. The prevalence of type 2 diabetes mellitus and hypertension were significantly higher in patients with SCS compared with those with nonfunctioning tumors. CONCLUSION: Functioning tumors, especially those with subclinical cortisol excess, are commonly found in patients with adrenal incidentalomas, although malignancy is rare. In addition, patients with SCS in adrenal incidentalomas have adverse metabolic and cardiovascular profiles.

Association between serum adipocytokine levels and microangiopathies in patients with type 2 diabetes mellitus.

AIMS/INTRODUCTION: It is thought that adipocytokines contribute to the increased risk of vascular complications in type 2 diabetes. However, there is still limited information on the relationship between microangiopathies and adipocytokines, such as adiponectin, leptin and tumor necrosis factor-α (TNF-α) in patients with type 2 diabetes. MATERIALS AND METHODS: The present study examined the relationship between fasting serum adiponectin, leptin, and TNF-α levels and microangiopathies in Korean type 2 diabetes. A total of 153 patients were recruited and evaluated for diabetic nephropathy, retinopathy and neuropathy. Serum adiponectin, TNF-α and leptin levels were measured. RESULTS: Serum adiponectin levels were significantly lower in patients with nephropathy than in those without nephropathy (P = 0.017), and were significantly higher in patients with retinopathy or neuropathy than those without retinopathy or neuropathy (P = 0.01 and P = 0.002, respectively). The mean levels of leptin were significantly higher in patients with neuropathy than in those without neuropathy (P = 0.002). The mean levels of TNF-α were not significantly different according to any of the three microangiopathies. Multivariate logistic regression analysis showed that the odds ratio for the presence of neuropathy in the highest tertile of adiponectin was 4.3 (95% confidence interval 1.59-11.62), as compared with the patients in the lowest tertile of adiponectin level. CONCLUSIONS: Levels of adipocytokines were significantly different according to the presence of each microangiopathy. In particular, higher serum adiponectin was independently associated with increased odds for the presence of neuropathy. Future prospective studies with larger numbers of patients are required to establish a direct relationship between plasma adipocytokine concentrations and the development or severity of diabetic microangiopathies.

Associations of serum fetuin-A levels with insulin resistance and vascular complications in patients with type 2 diabetes.

OBJECTIVE: We examined the relationship between serum fetuin-A, insulin resistance (IR), metabolic syndrome (MS) and vascular complications including cardiac autonomic neuropathy (CAN) in patients with type 2 diabetes mellitus (T2DM). METHODS: A total of 172 T2DM patients were recruited and evaluated for diabetic microangiopathies (nephropathy, retinopathy and peripheral neuropathy) including CAN. Serum fetuin-A levels were measured by enzyme-linked immunosorbent assay (ELISA), and the IR was assessed by the index of homeostasis model [homeostasis model assessment-insulin resistance (HOMA-IR)]. Atherosclerotic burden was assessed by ankle-brachial index (ABI) and brachial-ankle pulse wave velocity (baPWV). RESULTS: Serum fetuin-A levels showed significant positive correlations with HOMA-IR (r = 0.196, p = 0.022), and the mean levels of HOMA-IR were significantly increased progressively across fetuin-A tertiles (p for trend = 0.044). Serum fetuin-A showed significant positive correlations with baPWV, systolic blood pressure (BP), total cholesterol, triglycerides, serum fasting c-peptide and negative correlations with ABI. Serum fetuin-A levels were also negatively correlated with serum adiponectin and positively correlated with serum tumour necrosis factor-α (TNF-α). The mean levels of serum fetuin-A were not significantly different according to the presence of each microangiopathies including CAN. Also, the mean levels of serum fetuin-A were not different between patients with MS and without MS. CONCLUSIONS: This present study showed that levels of serum fetuin-A are significantly associated with IR and arterial stiffness assessed by baPWV, while there are no associations with each microangiopathies in patients with T2DM.

Lung nodule detected by F-18 fluorodeoxyglucose positron emission tomography-computed tomography in patients with papillary thyroid cancer, negative 131I whole body scan, and undetectable serum-stimulated thyroglobulin levels: two case reports.

INTRODUCTION: When a pulmonary nodular lesion is detected by F-18 fluorodeoxyglucose positron emission tomography-computed tomography in a patient with post-surgical papillary thyroid carcinoma with undetectable serum-stimulated thyroglobulin levels and negative 131I whole body scan, diagnosis and management of the nodule may be confusing. CASE PRESENTATION: We describe two post-surgical patients with papillary thyroid carcinoma who showed pulmonary nodular lesions detected by F-18 fluorodeoxyglucose positron emission tomography-computed tomography. In both cases serum-stimulated thyroglobulin levels were undetectable and nodular lesions were not detected by 131I whole body scan. In the first case, a 64-year-old Asian woman showed one focal increased fluorodeoxyglucose uptake lesion in the right lower lobe of one of her lungs. Based on the histologic study, the pulmonary nodular lesion was diagnosed as a solitary pulmonary metastasis from papillary thyroid carcinoma. In the second case, a 59-year-old Asian woman showed a new pulmonary nodule in the right lower lobe. The computed tomography scan of her chest revealed a 9mm nodule in the anterior basal segment and another tiny nodule in the posterior basal segment of the right lower lobe. Six months later, both nodules had increased in size and miliary disseminated nodules were also seen in both lungs. Based on their histology, the pulmonary nodular lesions were considered to be primary lung adenocarcinoma. CONCLUSIONS: The present cases emphasize that physicians should be cautious and make efforts for an accurate diagnosis of pulmonary nodules detected on F-18 fluorodeoxyglucose positron emission tomography-computed tomography in patients with papillary thyroid carcinoma with no evidence of metastasis such as negative 131I whole body scan and undetectable stimulated serum thyroglobulin levels.

Impact of clinicopathologic factors on subclinical central lymph node metastasis in papillary thyroid microcarcinoma.

PURPOSE: We evaluated whether the clinicopathological factors of papillary thyroid microcarcinoma (PTMC), especially tumoe size, are associated with subcinical central lymph node metastasis. MATERIALS AND METHODS: A total of 160 patients diagnosed with PTMC who underwent total thyroidectomy with bilateral central lymph node dissection were enrolled in this study. All patients were clinically lymph node negative PTMC. Patients were divided into 2 groups according to the size of tumor (≤5 mm vs. >5 mm). Clinicopathologic risk factors for subclinical central lymph node metastasis were analyzed. RESULTS: Subclinical central lymph node metastasis was detected in 61 (38.1%). Patients with tumors ≤5 mm had a lower frequency of extrathyroidal extension, multifocality and subclinical central lymph node metastasis. On multivariate analysis, only male and tumor size >5 mm were independent predictors of subclinical central lymph node metastasis; age, multifocality, bilaterality, extrathyroidal extension, lymphvascular invasion and lymphocytic thyroiditis were not. CONCLUSION: In this study, male and tumor size >5 mm were two independent predictive factors for subclinical central lymph node metastasis in PTMC. These are easier factors to assess before surgery than other factors when planning the central lymph node dissection. However, further long-term follow-up studies are needed to confirm the prognostic significance of subclinical central lymph node metastasis in PTMC.

Association of serum adipocytokine levels with cardiac autonomic neuropathy in type 2 diabetic patients.

BACKGROUND: Cardiac autonomic neuropathy (CAN) is a common complication of diabetes associated with poor prognosis. In addition, the autonomic imbalance is associated with cardiovascular disease (CVD) in diabetes. It is thought that adipocytokines contribute to the increased risk of vascular complications in patients with type 2 diabetes mellitus (T2DM). However, literature data on the association between CAN with adipocytokines such as leptin, tumor necrosis factor-alpha (TNF-alpha), adiponectin in subjects with T2DM is limited.Therefore, in the present study, we examined the relationship between fasting serum leptin, TNF- alpha and adiponectin and CAN in Korean T2DM patients. METHODS: A total of 142 T2DM patients (94 males, 48 females) were recruited. CAN was assessed by the five tests according to the Ewing's protocol and the time and frequency domain of the heart rate variability (HRV) was evaluated. Serum TNF-alpha and adiponectin levels were measured using enzyme-linked immunosorbent assay and serum leptin levels were measured using radioimmunoassay. RESULTS: Although, the mean levels of leptin, TNF-alpha and adiponectin were not significantly different between the groups with and without CAN, the levels of leptin and adiponectin had a tendency to increase as the score of CAN increased (p = 0.05, p = 0.036). Serum leptin levels demonstrated a negative correlation with low frequency (LF) in the upright position (p = 0.037). Regarding TNF-alpha, a significant negative correlation was observed with SDNN and RMSSD in the upright position (p = 0.023, p = 0.019). Adiponectin levels were not related to any HRV parameters. Multivariate logistic regression analysis demonstrated that the odds of CAN increased with a longer duration of diabetes (1.25, [1.07-1.47]) and higher homeostatic model of assessment-insulin resistance (HOMA-IR) (5.47, [1.8-16.5]). The relative risks for the presence of CAN were 14.1 and 51.6 for the adiponectin 2nd, 3rd tertiles when compared with first tertile (p-value for trend = 0.022). CONCLUSIONS: In the present study, the higher serum adiponectin levels and HOMA-IR were associated with an increased risk for the presence of CAN. Also, the CAN score correlated with the serum adiponectin. Serum adipocytokines such as leptin and TNF-alpha were significantly correlated with parameters of HRV, representative markers of CAN. Future prospective studies with larger number of patients are required to establish a direct relationship between plasma adipocytokine concentrations and the development or severity of CAN.

p53-Reactivating small molecules induce apoptosis and enhance chemotherapeutic cytotoxicity in head and neck squamous cell carcinoma.

We evaluate whether p53-reactivating (p53RA) small molecules induce p53-dependent apoptosis in head and neck squamous cell carcinoma (HNSCC), a question that has not been previously addressed in head and neck cancer. PRIMA-1, CP-31398, RITA, and nutlin-3 were tested in four human HNSCC cell lines differing in TP53 status. Cell growth, viability, cell cycle progression, and apoptosis after treatment with p53RA small molecules individually or in combination with chemotherapeutic agents were assessed. Prominent p53 reactivation was observed in mutant TP53-bearing tumor cell lines treated with PRIMA-1 or CP-31398, and in wild-type TP53-bearing cell lines treated with nutlin-3. Cell-cycle arrest and apoptosis induced by p53RA small molecules were associated with upregulation of p21 and BAX, and cleavage of caspase-3. Nutlin-3 showed maximal growth suppression in tumor cells showing MDM2-dependent p53 degradation. High-dose treatment with p53RA small molecules also induced apoptosis in cell lines independent of p53 or MDM2 expression. In combination therapy, p53RA small molecules enhanced the anti-tumor activity of cisplatin, 5-fluorouracil, paclitaxel, and erlotinib against HNSCC cells. The p53RA small molecules effectively restored p53 tumor-suppressive function in HNSCCs with mutant or wild-type TP53. The p53RA agents may be clinically useful against HNSCC, in combination with chemotherapeutic drugs.

Disialoganglioside GD3 synthase expression recruits membrane transglutaminase 2 during erythroid differentiation of the human chronic myelogenous leukemia K562 cells.

By employing proteomics analysis tool, we examined the effects of GD3 synthase expression on the differentiation properties of chronic myelogenous leukemia (CML)-derived leukemia cells K562. Forced expression of GD3 synthase induced erythroid differentiation as determined by an increase in glycophorin A expression and synthesis of hemoglobins. The proteomic analysis revealed that 15 proteins were increased by GD3 synthase. In contrast, we observed three protein gel spots decreased in contents in the cell membranes of GD3 synthase-transfected K562 cells. Among the increased proteins, membrane transglutaminase 2 (TG2) was specifically increased in the cell membrane of GD3 synthase-transfected K562 cells. Then, we generated the GD3 synthase-transfected cells in the K562 cells. Interestingly, the TG2 level was increased in GD3 synthase-transfected cells compared with vector- and plasma membrane-associated ganglioside sialidase (Neu3)-transfected cells. In addition, its ability to be photoaffinity-labeled with [alpha-(32)P]GTP was also increased in the GD3 synthase- and TG2-transfected cells. Moreover, small interfering RNA (siRNA) analysis for the GD3 synthase showed the decrease or abolishment of the membrane TG2. Finally, GD3 synthase-transfected cells accelerated the erythroid differentiation. Therefore, we propose that the recruitment of TG2 into membranes by GD3 might play an important role in the erythroid differentiation in K562 cells.

Human AQP5 plays a role in the progression of chronic myelogenous leukemia (CML).

Aquaporins (AQPs) have previously been associated with increased expression in solid tumors. However, its expression in hematologic malignancies including CML has not been described yet. Here, we report the expression of AQP5 in CML cells by RT-PCR and immunohistochemistry. While normal bone marrow biopsy samples (n = 5) showed no expression of AQP5, 32% of CML patient samples (n = 41) demonstrated AQP5 expression. In addition, AQP5 expression level increased with the emergence of imatinib mesylate resistance in paired samples (p = 0.047). We have found that the overexpression of AQP5 in K562 cells resulted in increased cell proliferation. In addition, small interfering RNA (siRNA) targeting AQP5 reduced the cell proliferation rate in both K562 and LAMA84 CML cells. Moreover, by immunoblotting and flow cytometry, we show that phosphorylation of BCR-ABL1 is increased in AQP5-overexpressing CML cells and decreased in AQP5 siRNA-treated CML cells. Interestingly, caspase9 activity increased in AQP5 siRNA-treated cells. Finally, FISH showed no evidence of AQP5 gene amplification in CML from bone marrow. In summary, we report for the first time that AQP5 is overexpressed in CML cells and plays a role in promoting cell proliferation and inhibiting apoptosis. Furthermore, our findings may provide the basis for a novel CML therapy targeting AQP5.

Role of human aquaporin 5 in colorectal carcinogenesis.

While overexpression of several aquaporins (AQPs) has been reported in different types of human cancer, the role of AQPs in carcinogenesis has not been clearly defined. Here, by immunochemistry, we have found expression of AQP5 protein in 62.8% (59/94) of resected colon cancer tissue samples as well as association of AQP5 with liver metastasis. We then demonstrated that overexpression of human AQP5 (hAQP5) induces cell proliferation in colon cancer cells. Overexpression of wild-type hAQP5 increased proliferation and phosphorylation of extracellular signal-regulated kinase-1/2 in HCT116 colon cancer cells whereas these phenomena in hAQP5 mutants (N185D and S156A) were diminished, indicating that both membrane association and serine/threonine phosphorylation of AQP5 are required for proper function. Interestingly, overexpression of AQP1 and AQP3 showed no differences in extracellular signal-regulated kinase-1/2 phosphorylation, suggesting that AQP5, unlike AQP1, may be involved in signal transduction. Moreover, hAQP5-overexpressing cells showed an increase in retinoblastoma protein phosphorylation through the formation of a nuclear complex with cyclin D1 and CDK4. Small interfering RNA analysis confirmed that hAQP5 activates the Ras signaling pathway. These data not only describe the induction of hAQP5 expression during colorectal carcinogenesis but also provide a molecular mechanism for colon cancer development through the interaction of hAQP5 with the Ras/extracellular signal-regulated kinase/retinoblastoma protein signaling pathway, identifying hAQP5 as a novel therapeutic target.

Expression of aquaporin 5 (AQP5) promotes tumor invasion in human non small cell lung cancer.

The aquaporins (AQP) are water channel proteins playing a major role in transcellular and transepithelial water movement. Recently, the role of AQPs in human carcinogenesis has become an area of great interest. Here, by immunohistochemistry (IHC), we have found an expression of AQP5 protein in 35.3% (IHC-score: > or = 1, 144/408) of the resected NSCLC tissue samples. Cases with AQP5-positive status (IHC-score: > or = 2) displayed a higher rate of tumor recurrence than negative ones in NSCLC (54.7% vs. 35.1%, p = 0.005) and worse disease-free survival (p = 0.033; OR = 1.52; 95%CI: 1.04-2.23). Further in vitro invasion assay using BEAS-2B and NIH3T3 cells stably transfected with overexpression constructs for full length wild-type AQP5 (AQP5) and its two mutants, N185D which blocks membrane trafficking and S156A which blocks phosphorylation on Ser156, showed that AQP5 induced cell invasions while both mutants did not. In BEAS-2B cells, the expression of AQP5 caused a spindle-like and fibroblastic morphologic change and losses of cell-cell contacts and cell polarity. Only cells with AQP5, not either of two mutants, exhibited a loss of epithelial cell markers and a gain of mesenchymal cell markers. In a human SH3-domains protein array, cellular extracts from BEAS-2B with AQP5 showed a robust binding activity to SH3-domains of the c-Src, Lyn, and Grap2 C-terminal. Furthermore, in immunoprecipitation assay, activated c-Src, phosphorylated on Tyr416, showed a stronger binding activity to cellular extracts from BEAS-2B with AQP5 compared with N185D or S156A mutant. Fluorescence in situ hybridization (FISH) analysis failed to show evidence of genomic amplification, suggesting AQP5 expression as a secondary event. Based on these clinical and molecular observations, we conclude that AQP5, through its phosphorylation on Ser156 and subsequent interaction with c-Src, plays an important role in NSCLC invasion and, therefore, may provide a unique opportunity for developing a novel therapeutic target as well as a prognostic marker in NSCLC.

Short-term changes in bone and mineral metabolism following gastrectomy in gastric cancer patients.

Changes in bone and mineral metabolism that occur after gastrectomy have long been recognized. Gastrectomy has been identified as a risk factor for decreased bone mass and the increased fracture incidence. Previous investigations concerning postgastrectomy bone disease have been observational studies. No prospective studies have been reported that quantify the amount of bone loss after gastrectomy within the same patients. This study investigated 46 patients undergoing gastrectomy for gastric adenocarcinoma and analyzed 36 patients (58.1+/-10.8 years, 24 men and 12 women) who had dual energy X-ray absorptiometry (DXA) performed before and 1 year after gastrectomy. Systemic adjuvant chemotherapy was administered to 14 patients. Blood was sampled from all patients to determine serum calcium, phosphorous, and bone turnover marker levels before gastrectomy and at 1, 3, 6 and 12 months after surgery and for serum parathyroid hormone (PTH) and 25-hydroxyvitamin D levels before and 12 months after surgery. The mean bone loss in the lumbar spine, total hip, femoral neck, and trochanter, which was calculated as the percentage change from the baseline to the level measured at 12 months, was 5.7% (P<0.01), 5.4% (P<0.01), 6.6% (P<0.01) and 8.7% (P<0.01), respectively. Bone loss was generally greater in the group receiving chemotherapy. The serum calcium and phosphorous levels were not changed significantly and remained within the normal range throughout the observation period. After gastrectomy, the level of ICTP increased and reached a peak at 1 and 3 months, and progressively declined to baseline by 12 months. The osteocalcin levels were not coupled to an increase before 6 months. The level of 25-hydroxyvitamin D at 12 months postgastrectomy was not significantly changed compared to the baseline, however, the PTH levels increased by a mean of 63.6% at 12 months compared to the baseline (P<0.01). Significant correlations were found between the percent change in the BMD at the lumbar spine and total hip and the percentage change for the PTH level from their baselines to 12 months. The changes in the BMD at total hip, femoral neck, and trochanter also correlated to the change in body weight at 12 months. The data obtained by this study provides evidence that profound bone loss occurs in the setting of a bone remodeling imbalance during the early postgastrectomy period and allows the speculation that the gastrectomy related bone loss may be partially due to an overproduction of PTH.

Clinical, histopathological, and molecular characteristics of papillary thyroid microcarcinoma.

BACKGROUND: Papillary thyroid microcarcinoma (PTMC) detection has been increasing explosively with the recent improvement and application of new diagnostic tools. However, the associations between clinical, histopathological, and molecular findings are often not fully evaluated in reaching a decision for the treatment of PTMC. OBJECTIVE: To retrospectively evaluate the patients diagnosed with PTMC in terms of the clinical and histopathological features and the immunohistochemical findings of specific molecular markers, and thereby provide an association with the disease prognosis for the Korean patients. DESIGN: We have reviewed the clinical records of patients who underwent surgery for thyroid cancer (during January 2000 to August 2006 interval) at St. Mary's Hospital, Seoul, Korea. The clinical and histopathological characteristics of 217 PTMCs were evaluated cross-sectionally after surgery, and immunohistochemical staining of p53, epidermal growth factor receptor (EGFR), Ki-67, and cyclooxygenase-2 (COX-2) for 87 isolated specimens was performed. MAIN OUTCOME: The proportions of extrathyroid extension, lymphatic invasion, and lymph node metastasis cited were 31.9%, 19.2%, and 17.8%, respectively. However, distant metastasis was not noted. Fifty-three (27.1%) patients had multiplicity, and 39 patients (20.0%) had bilateral disease. A primary tumor larger than 5 mm was significantly associated with extrathyroid extension, lymphatic invasion, and lymph node metastasis. The absence of EGFR expression shown by immunohistochemical analysis was closely correlated with extrathyroid extension and lymph node metastasis, while the absence of COX-2 expression was associated with multiplicity and bilaterality. CONCLUSION: Many patients with PTMC had clinical and histopathological poor prognostic factors. The expression of molecular markers, such as EGFR and COX-2, may have a role in the prognosis of PTMC. When considering all of the prognostic factors, a more tailored management approach appears to be necessary for patients with PTMC.

Acromegaly associated with type 2 diabetes showing normal IGF-1 levels under poorly controlled glycemia.

Acromegaly is caused by excessive secretion of growth hormone (GH), and a resultant persistent elevation of insulin-like growth factor-1 (IGF-1) levels. Diabetes mellitus is accompanied in some acromegalic patients with insulin resistance. We encountered a type-2 diabetic patient who had a poorly controlled glycemic state and was diagnosed as acromegaly with normal IGF-1 levels. The patient showed definite acromegalic features. However, in the first screening test, GH levels were high and IGF-1 levels were inappropriately normal so the results were not close to the diagnosis of acromegaly. After moderate glycemic control, an oral glucose suppression test was performed, showing no suppressed GH response. TRH test revealed paradoxical increases in growth hormone levels and a brain MRI discovered a pituitary adenoma. After several-months insulin treatment, IGF-1 levels were increased to the abnormal state and GH levels were decreased without treatment for acromegaly. Here we report the rare case of acromegaly that presents inappropriately normal IGF-1 levels at the time of diagnosis in uncontrolled type 2 diabetic patient and shows increased IGF-1 levels after glycemic control with insulin therapy. When evaluating acromegaly in type 2 diabetes under poorly controlled glycemia, cautious IGF-1 analysis is needed after sufficient glycemic control.