Low Molecular Weight Protein Tyrosine Phosphatase Expression Research Articles

Violacein switches off low molecular weight tyrosine phosphatase and rewires mitochondria in colorectal cancer cells

In the last decade, emerging evidence has shown that low molecular weight protein tyrosine phosphatase (LMWPTP) not only contributes to the progression of cancer but is associated with prostate low survival rate and colorectal cancer metastasis. We report that LMWPTP favors the glycolytic profile in some tumors. Therefore, the focus of the present study was to identify metabolic enzymes that correlate with LMWPTP expression in patient samples. Exploratory data analysis from RNA-seq, proteomics, and histology staining, confirmed the higher expression of LMWPTP in CRC. Our descriptive statistical analyses indicate a positive expression correlation between LMWPTP and energy metabolism enzymes such as acetyl-CoA carboxylase (ACC) and fatty acid synthase (FASN). In addition, we examine the potential of violacein to reprogram energetic metabolism and LMWPTP activity. Violacein treatment induced a shift of glycolytic to oxidative metabolism associated with alteration in mitochondrial efficiency, as indicated by higher oxygen consumption rate. Particularly, violacein treated cells displayed higher proton leak and ATP-linked oxygen consumption rate (OCR) as an indicator of the OXPHOS preference. Notably, violacein is able to bind and inhibit LMWPTP. Since the LMWPTP acts as a hub of signaling pathways that offer tumor cells invasive advantages, such as survival and the ability to migrate, our findings highlight an unexplored potential of violacein in circumventing the metabolic plasticity of tumor cells.

Platelet-dependent signaling and Low Molecular Weight Protein Tyrosine Phosphatase expression promote aggressive phenotypic changes in gastrointestinal cancer cells

Over the last decades, some members of the protein tyrosine phosphatase family have emerged as cancer promoters. Among them, the Low Molecular Weight Protein Tyrosine Phosphatase (LMWPTP) has been described to be associated with colorectal cancer liver metastasis and poor prostate cancer prognosis. Of importance in the process of cancer progression and metastasis is the interaction between tumor cells and platelets, as the latter are thought to promote several tumor hallmarks. Here, we examine to what extent LMWPTP expression in tumor cells affects their interaction with platelets. We demonstrate that the gene encoding LMWPTP is overexpressed in upper gastrointestinal (GI) cancer cell as well as colorectal cancer, and subsequently employ cell line models to show that the level of this phosphatase may be further augmented in the presence of platelets. We demonstrate that tumor-platelet interaction promotes GI tumor cell proliferation. Additionally, using know-down/-out models we show that LMWPTP expression in cancer cells contributes to a more efficient interaction with platelets and drives platelet-induced proliferation. These data are the first to demonstrate that phosphatases play a positive role in the tumor-promoting activities of platelets, with LMWPTP emerging as a key player promoting oncogenic phenotypic changes in tumor cells.

Prediction of Time to Castration-Resistant Prostate Cancer Using Low-Molecular-Weight Protein Tyrosine Phosphatase Expression for Men with Metastatic Hormone-Naïve Prostate Cancer

Introduction: Low-molecular-weight protein tyrosine phosphatase (LMW-PTP) expression affects carcinogenesis in various cancers and has been associated with determining the overall survival among men with metastatic hormone-naïve prostate cancer (mHNPC). In this study, we analyzed the value of LMWPTP for prediction of time to castration-resistant prostate cancer (CRPC) for men with mHNPC. Materials and Methods: We retrospectively enrolled 45 men with mHNPC who were diagnosed from 2003 to 2009. All patients had received androgen deprivation therapy as first-line treatment. Prostate cancer tissues (pre-treatment needle biopsies) were immunohistochemically stained for LMW-PTP. Multivariate analyses (Cox proportional hazard model) were used to correlate baseline clinical factors of age, prostate-specific antigen (PSA), Gleason scores, T stage, N stage, extent of disease on bone scan (EOD), LMW-PTP expression and time to CRPC. Continuous variables were classified as dichotomous. Results: Median age and PSA were 70.0 years and 87.8 ng/mL respectively. Median time to CRPC was 40.2 months. Median time to CRPC was significantly shorter in the high LMW-PTP group (14.8 months) than that in the low LMW-PTP group (86.3 months, p < 0.01). In multivariate analysis, age ≥70 years and high LMW-PTP expression were significant predictors of time to CRPC.

Abstract 1546: Low-molecular-weight protein tyrosine phosphatase is a useful biomarker for predicting postoperative biochemical recurrence in RM0 prostate cancer

Abstract Objective: Low-molecular-weight protein tyrosine phosphatase (LMPTP) has a molecular weight of 18 kDa. Studies have shown that the elevated expression of this enzyme is associated with poor prognosis of certain cancers. For prostate cancer, positive surgical margins are considered an important predictor of such recurrences. However, biochemical recurrence is observed even with a negative surgical margin (RM0), and some cases require salvage therapy. Few reports have identified useful biomarkers to predict these cases. In this study, we investigated whether LMPTP expression level could be a biomarker of these recurrences in RM0 prostate cancer. Methods: The subjects were 119 patients who underwent a total prostatectomy with a negative resection margin, at our hospital, between 2007 and 2011. Immunostaining of 2 TMA samples per patient was performed to evaluate LMPTP expression levels. Expression was classified as negative, 0; weak, 1; moderate, 2; or strong, 3 by two pathologists. Each patient was given a total expression score consisting of 13 levels from 0 to 12. By constructing a ROC curve, LMPTP expressions were categorized into either a high expression group (score: ≥5) or a low expression group (score: ≤4). We then examined the relationship of LMPTP expression level with other pathological observations and biochemical recurrence. Cancer survival analysis was performed using the Kaplan-Meier method, log-rank test, and Cox's proportional hazards model. The threshold for statistical significance was set at p &amp;lt; 0.05. Results: Evaluation of immunostained samples by the two pathologists were highly reliable, with an Intraclass correlation (ICC) score for two distinct measurements of 0.77 and 0.98, respectively. 68 patients (57.1%) were categorized in the LMPTP high expression group; and 51 patients (42.9%), in the low expression group. The log-rank test revealed that early biochemical recurrence was observed in the high LMPTP expression group (p = 0.0011). In addition, both the univariate and multivariate analyses revealed that high LMPTP expression level was an independent prognostic factor for biochemical recurrence. Furthermore, in the RM0 cases, high LMPTP expression level (HR:3.62, p = 0.023) was shown to be a better prognostic factor than Gleason score (GS; p = 0.406). Lastly, high LMPTP expression level strongly correlated with the Ki-67 proliferation marker (p &amp;lt; 0.0001), suggesting that LMPTP is expressed in tumor cells with high proliferative capacity. Conclusion: This study suggests that immunostaining of LMPTP is a better predictor of biochemical recurrence than GS in cases of RM0 total prostatectomy. This biomarker can be evaluated with a single immunostaining with high reproducibility, and can therefore be a clinically applicable tool in postoperative follow-ups and as a predictor of recurrence that require salvage therapy. Citation Format: Hirofumi Kurose, Kosuke Ueda, Reiichiro Kondo, Kiyoaki Nishihara, Makoto Nakiri, Mitsunori Matsuo, Shigetaka Suekane, Sakiko Sanada, Sachiko Ogasawara, Tsukasa Igawa, Hirohisa Yano. Low-molecular-weight protein tyrosine phosphatase is a useful biomarker for predicting postoperative biochemical recurrence in RM0 prostate cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1546.

Evaluation of ability of low-molecular-weight protein tyrosine phosphatase expression to predict primary hormone therapy efficacy among men with metastatic hormone-naïve prostate cancer.

316 Background: Protein tyrosine phosphorylation is an important event associated with the processes of cellular metabolism and is regulated by protein tyrosine kinases and protein tyrosine phosphatases (PTPs). Recent studies showed that low-molecular-weight protein tyrosine phosphatase (LMW-PTP) could be a biomarker for predicting overall survival among men with metastatic hormone-naïve prostate cancer (mHNPC). In this study, we evaluated the value of LMW-PTP expression for predicting primary hormone therapy efficacy among men with mHNPC. Methods: A total of 45 men with mHNPC diagnosed from 2003 to 2009 were enrolled in this retrospective study. All patients had received androgen ablation therapy as first-line treatment. Prostate cancer tissues obtained by needle biopsies before treatment were immunohistochemically stained for LMW-PTP. Correlations between the baseline clinical factors of age, PSA, Gleason scores, T stage, N stage, extent of disease on bone scan (EOD), LMW-PTP expression and primary hormone therapy efficacy (time to castration-resistant prostate cancer [CRPC]) were investigated by multivariate analysis using the Cox proportional hazard model. Continuous variables were classified as dichotomous. Results: Median age and PSA were 70.0 years and 87.8 ng/mL, respectively. Distribution number of Gleason score 8-10/6-7, T4/≤3, N1/0, and EOD 3-4/0-2 were 30 (66.7%)/15 (33.3%), 13 (28.9%)/32 (71.1%), 25 (55.6%)/20 (44.4%), and 7 (15.5%)/38 (84.5%). Median time to CRPC was 40.2 months. Patients with high LMW-PTP expression had significantly shorter time to CRPC than those with low expression (p = 0.005). Multivariate analysis showed that age (≥70 vs &lt; 70, HR 2.5, 95%CI 0.96-6.32, p = 0.060) and LMW-PTP expression (high vs low; HR 3.3, 95%CI 1.2-8.6, p = 0.02) were significant prognostic biomarkers for predicting time to CRPC. Conclusions: LMW-PTP expression could be a potential biomarker for predicting primary hormone therapy efficacy among men with mHNPC.

Low-molecular-weight protein tyrosine phosphatase expression as a prognostic factor for men with metastatic hormone-naïve prostate cancer

ObjectivesRecent studies have demonstrated that up-front docetaxel combined with androgen deprivation therapy (ADT) prolongs survival in some patients with metastatic hormone-naïve prostate cancer (mHNPC). However, new biomarkers for selecting personalized treatment strategies for mHNPC are warranted. We evaluated the value of low-molecular-weight protein tyrosine phosphatase (LMW-PTP) expression as a prognosticator in men with mHNPC. Methods and materialsA total of 48 men with mHNPC diagnosed from 2003 to 2009 were enrolled in this study. Prostate cancer tissues obtained by needle biopsies were immunohistochemically stained for LMW-PTP. Correlations between LMW-PTP expression and clinicopathological characteristics were then assessed. ResultsAt the time of analysis, 29 (60.4%) patients were alive, whereas 15 (31.3%) and 4 (8.3%) died of prostate cancer and nonprostate cancer, respectively. Of these, 29 (60.4%) had low LMW-PTP expression and 19 (39.6%) had high expression. Median overall survival (OS) for patients with high LMW-PTP expression was not reached and that for patients with low LMW-PTP expression was 23.8 months. High LMW-PTP expression was significantly correlated with a shorter OS compared with low LMW-PTP expression (P = 0.01). Moreover, multivariate analysis showed that Gleason score (≥8 vs.≤7; HR = 5.8, 95% CI: 1.3–26.5, P = 0.02) and LMW-PTP expression (high vs. low; HR = 2.7, 95% CI: 1.0–7.2, P = 0.04) were independent prognostic factors for OS. ConclusionsLMW-PTP is a potential biomarker to predict OS in patients with mHNPC.

The usefullness of low-molecular-weight protein tyrosine phosphatase as a prognostic factor in metastatic hormone-sensitive prostate cancer.

277 Background: Protein tyrosine phosphorylation is an important event that is associated with the processes of cellular metabolism. The control of protein tyrosine phosphorylation is regulated by protein tyrosine kinases and protein tyrosine phosphatases (PTP). Thus, these proteins can control diverse processes like cell proliferation, adhesion, and migration. Recent studies revealed the importance of PTP in the oncogenic process, especially, the significance of low-molecular-weight protein tyrosine phosphatase (LMW-PTP) as a prognostic factor in early prostate cancer. In this study, we evaluated the significance of expression of LMW-PTP in metastatic hormone-sensitive prostate cancer (mHSPC) as a prognostic biomarker. Methods: We identified consecutive 48 men with mHSPC. The prostate cancer tissues obtained by needle biopsies were immunohistochemically stained for LMW-PTP. A pathologist blindly evaluated the expression of LMW-PTP and intensity of staining was classified into two groups, high (diffusely strongly positive) and low (at least focally weak-negative). The correlations between clinicopathological characteristics including age, initial PSA levels, Gleason scores, T stage, N stage, extend of disease on bone scan (EOD), and expression of LMW-PTP and overall survival (OS) were analyzed by multivariate analysis using cox proportional hazards regression model. Results: Median age was 70.0 years and median initial PSA level was 85.4 ng/mL. Gleason scores were 6-7 in 16 patients (33.4%) and 8-10 in 32 patients (66.6%). Clinical T stages were T2-3 disease in 34 patients (70.8%) and T4 in 14 patients (29.2%). Clinical N stages were N0 disease in 21 patients (43.7%) and N1 in 27 patients (56.3%). EOD grades were 0 in 13 patients (27.0%), 1 in 12 patients (25.0%), 2 in 14 patients (29.2%), 3 in 8 patients (16.7%), and 4 in 1 patient (2.1%). Multivariate analysis revealed that Gleason score (≥ 8 vs ≤ 7; HR: 5.795, 95%CI: 1.266-26.520, p = 0.024) and LMW-PTP expression (high vs low; HR: 2.736, 95%CI: 1.035-7.232, p = 0.042) were independent prognostic factors for OS. Conclusions: LMW-PTP might be a potential biomarker to predict overall survival for men with mHSPC.

Deletion of low molecular weight protein tyrosine phosphatase (Acp1) protects against stress-induced cardiomyopathy.

The low molecular weight protein tyrosine phosphatase (LMPTP), encoded by the ACP1 gene, is a ubiquitously expressed phosphatase whose in vivo function in the heart and in cardiac diseases remains unknown. To investigate the in vivo role of LMPTP in cardiac function, we generated mice with genetic inactivation of the Acp1 locus and studied their response to long‐term pressure overload. Acp1−/− mice develop normally and ageing mice do not show pathology in major tissues under basal conditions. However, Acp1−/− mice are strikingly resistant to pressure overload hypertrophy and heart failure. Lmptp expression is high in the embryonic mouse heart, decreased in the postnatal stage, and increased in the adult mouse failing heart. We also show that LMPTP expression increases in end‐stage heart failure in humans. Consistent with their protected phenotype, Acp1−/− mice subjected to pressure overload hypertrophy have attenuated fibrosis and decreased expression of fibrotic genes. Transcriptional profiling and analysis of molecular signalling show that the resistance of Acp1−/− mice to pathological cardiac stress correlates with marginal re‐expression of fetal cardiac genes, increased insulin receptor beta phosphorylation, as well as PKA and ephrin receptor expression, and inactivation of the CaMKIIδ pathway. Our data show that ablation of Lmptp inhibits pathological cardiac remodelling and suggest that inhibition of LMPTP may be of therapeutic relevance for the treatment of human heart failure. © 2015 The Authors. The Journal of Pathology published by John Wiley & Sons Ltd on behalf of Pathological Society of Great Britain and Ireland.

Low-Molecular-Weight Protein Tyrosine Phosphatase Predicts Prostate Cancer Outcome by Increasing the Metastatic Potential

BackgroundLow-risk patients suffering from prostate cancer (PCa) are currently placed under active surveillance rather than undergoing radical prostatectomy. However, clear parameters for selecting the right patient for each strategy are not available, and new biomarkers and treatment modalities are needed. Low-molecular-weight protein tyrosine phosphatase (LMWPTP) could present such a target. ObjectiveTo correlate expression levels of LMWPTP in primary PCa to clinical outcome, and determine the role of LMWPTP in prostate tumor cell biology. Design, setting, and participantsAcid phosphatase 1, soluble (ACP1) expression was analyzed on microarray data sets, which were subsequently used in Ingenuity Pathway Analysis. Immunohistochemistry was performed on a tissue microarray containing material of 481 PCa patients whose clinicopathologic data were recorded. PCa cell line models were used to investigate the role of LMWPTP in cell proliferation, migration, adhesion, and anoikis resistance. Outcome measurements and statistical analysisThe association between LMWPTP expression and clinical and pathologic outcomes was calculated using chi-square correlations and multivariable Cox regression analysis. Functional consequences of LMWPTP overexpression or downregulation were determined using migration and adhesion assays, confocal microscopy, Western blotting, and proliferation assays. Results and limitationsLMWPTP expression was significantly increased in human PCa and correlated with earlier recurrence of disease (hazard ratio [HR]:1.99; p<0.001) and reduced patient survival (HR: 1.53; p=0.04). Unbiased Ingenuity analysis comparing cancer and normal prostate suggests migratory propensities in PCa. Indeed, overexpression of LMWPTP increases PCa cell migration, anoikis resistance, and reduces activation of focal adhesion kinase/paxillin, corresponding to decreased adherence. ConclusionsOverexpression of LMWPTP in PCa confers a malignant phenotype with worse clinical outcome. Prospective follow-up should determine the clinical potential of LMWPTP overexpression. Patient summaryThese findings implicate low-molecular-weight protein tyrosine phosphatase as a novel oncogene in prostate cancer and could offer the possibility of using this protein as biomarker or target for treatment of this disease.

Low molecular weight protein tyrosine phosphatase (LMWPTP) upregulation mediates malignant potential in colorectal cancer.

Phosphatases have long been regarded as tumor suppressors, however there is emerging evidence for a tumor initiating role for some phosphatases in several forms of cancer. Low Molecular Weight Protein Tyrosine Phosphatase (LMWPTP; acid phosphatase 1 [ACP1]) is an 18 kDa enzyme that influences the phosphorylation of signaling pathway mediators involved in cancer and is thus postulated to be a tumor-promoting enzyme, but neither unequivocal clinical evidence nor convincing mechanistic actions for a role of LMWPTP have been identified. In the present study, we show that LMWPTP expression is not only significantly increased in colorectal cancer (CRC), but also follows a step-wise increase in different levels of dysplasia. Chemical inhibition of LMWPTP significantly reduces CRC growth. Furthermore, downregulation of LMWPTP in CRC leads to a reduced migration ability in both 2D- and 3D-migration assays, and sensitizes tumor cells to the chemotherapeutic agent 5-FU. In conclusion, this study shows that LMWPTP is not only overexpressed in colorectal cancer, but it is correlated with the malignant potential of this cancer, suggesting that this phosphatase may act as a predictive biomaker of CRC stage and represents a rational novel target in the treatment of this disease.

Osteoblast Adhesion Dynamics: A Possible Role for ROS and LMW‐PTP

Reactive oxygen species (ROS) modulate a variety of intracellular events, but their role in osteoblast adhesion and spreading remains unclear. ROS is a very-known physiological modulators of Protein Tyrosine Phosphatases activities, mainly to low molecular weight protein tyrosine phosphatase (LMW-PTP) activity. As this biological mechanism is not clear in osteoblast adhesion, we decided to investigate ROS levels and phosphorylations of FAK and Src, identifying these proteins as potential substrates to LMW-PTP activity. Our results showed that during osteoblast adhesion/spreading (30 min and 2 h of seeding) the intracellular ROS content (hydrogen peroxide) is finely regulated by an effective anti-oxidant system [catalase and Superoxide Dismutase (SOD) activities were evaluated]. During the first 30 min of adhesion, there was an increase in ROS production and a concomitant increase in focal adhesion kinase (FAK) activity after its phosphorylation at Tyrosine 397 (Y397 ). Moreover, after 2 h there was a decrease in ROS content and FAK phosphorylation. There was no significant change in LMW-PTP expression at 30 min or 2 h. In order to validate our hypothesis that LMW-PTP is able to control FAK activity by modulating its phosphorylation status, we decided to overexpress and silence LMW-PTP in this context. Our results showed that FAK phosphorylation at Y397 was increased and decreased in osteoblasts with silenced or overexpressed LMW-PTP, respectively. Together, these data show that ROS modulate FAK phosphorylation by an indirect way, suggesting that a LMW-PTP/FAK supra-molecular complex is involved in transient responses during osteoblast adhesion and spreading.

S-Glutathionylation of LMW-PTP regulates VEGF-mediated FAK activation and endothelial cell migration

Although promising, the ability to regulate angiogenesis through delivery of VEGF remains an unrealized goal. We have shown previously that physiological levels of peroxynitrite (1 µM) are required for a VEGF-mediated angiogenic response, yet the redox-regulated mechanisms that govern the VEGF signal remain unexplored. We assessed the impact of VEGF and peroxynitrite on modifying redox-state, the level of reduced-glutathione (GSH) and S-glutathionylation on regulation of the low molecular weight protein tyrosine phosphatase (LMW-PTP) and focal adhesion kinase (FAK), which are key mediators of VEGF-mediated cell migration. Stimulation of human microvascular endothelial (HME) cells with VEGF (20 ng/ml) or peroxynitrite (1 µM) caused an immediate and reversible negative-shift in the cellular redox-state and thiol oxidation of LMW-PTP, which culminated in cell migration. VEGF causes reversible S-glutathionylation of LMW-PTP, which inhibits its phosphorylation and activity, and causes the transient activation of FAK. Modulating the redox-state using decomposing peroxynitrite (FeTPPS, 2.5 µM) or the GSH-precursor [N-acetylcysteine (NAC), 1 mM] caused a positive-shift of the redox-state and prevented VEGF-mediated S-glutathionylation and oxidative inhibition of LMW-PTP. NAC and FeTPPS prevented the activation of FAK, its association with LMW-PTP and cell migration. Inhibiting LMW-PTP expression markedly enhanced FAK activation and cell migration. Although mild oxidative stress achieved by combining VEGF with 0.1-0.2 mM peroxynitrite augmented cell migration, an acute shift to oxidative stress achieved by combining VEGF with 0.5 mM peroxynitrite induced and sustained FAK activation, and LMW-PTP S-glutathionylation, resulting in LMW-PTP inactivation and inhibited cell migration. In conclusion, our findings demonstrate that a balanced redox-state is required for VEGF to facilitate reversible S-glutathionylation of LMW-PTP, FAK activation and endothelial cell migration. Shifting the redox-state to reductive stress or oxidative stress inhibited the VEGF-mediated angiogenic response.

A possible mechanism of low molecular weight protein tyrosine phosphatase (LMW-PTP) activity modulation by glutathione action during human osteoblast differentiation

Objective Low molecular weight protein tyrosine phosphatases (LMW-PTPs) are a family of enzymes strongly involved in the regulation of cell growth and differentiation. Since there is no information concerning the relationship between osteoblastic differentiation and LMW-PTP expression/activity, we investigated its involvement during human osteoblast-like cells (hFOB 1.19) differentiation. It is known that LMW-PTP is regulated by an elegant redox mechanism, so we also observed how the osteoblastic differentiation affected the reduced glutathione levels. Design hFOB 1.19 cells were cultured in DMEM/F12 up to 35 days. The osteoblast phenotype acquisition was monitored by measuring alkaline phosphatase activity and mineralized nodule formation by Von Kossa staining. LMW-PTP activity and expression were measured using the p-nitrophenylphosphate as substrate and Western blotting respectively. Crystal violet assay determined the cell number in each experimental point. Glutathione level was determined by both HPLC and DNTB assays. Results LMW-PTP modulation was coincident with the osteoblastic differentiation biomarkers, such as alkaline phosphatase activity and presence of nodules of mineralization in vitro. Likewise LMW-PTP, the reduced glutathione-dependent microenvironment was modulated during osteoblastic differentiation. During this process, LMW-PTP expression/activity, as well as alkaline phosphatase and glutathione increased progressively up to the 21st day ( p < 0.001) of culturing, decreasing thereafter. Conclusions Our results clearly suggest that LMW-PTP expression/activity was rigorously modulated during osteoblastic differentiation, possibly in response to the redox status of the cells, since it seems to depend on suitable levels of reduced glutathione. In this way, we pointed out LMW-PTP as an important signaling molecule in osteoblast biology and bone formation.

The expression of low molecular weight protein tyrosine phosphatase is up‐regulated in 1,2‐dimethylhydrazine‐induced colon tumours in rats

Recent studies have assessed the role of low molecular weight protein tyrosine phosphatase (LMW-PTP) in cell transformation and tumour onset and progression, observing a significant increase in the expression of LMW-PTP mRNA and protein in human breast, colon, bladder and kidney tumour samples. Moreover, its enhanced expression is generally prognostic of a more aggressive cancer. To better understand the role of this protein during colon carcinogenesis and to study whether its overexpression is also observed in earlier phases of carcinogenesis, we studied its expression in colon tumours, induced in rats by treatment with 1,2-dimethylhydrazine (DMH), an animal model that resemble the sequential formation of histopathological lesions of spontaneous carcinogenesis in humans. The results show a significant increase in LMW-PTP expression in adenocarcinomas, suggesting that this phenomenon is associated with the onset of malignancy. Moreover a significant overexpression of LMW-PTP transcript is associated with tumours originating in the proximal (right) part of the colon, confirming an observation already reported for human colon cancer.

Modulation of Src Activity by Low Molecular Weight Protein Tyrosine Phosphatase During Osteoblast Differentiation

Background: Src kinase plays a critical role in bone metabolism, particularly in osteoclasts. However, the ability of Src kinase to modulate the activity of other bone cells is less well understood. In this work, we examined the expression and activity of Src and low molecular weight protein tyrosine phosphatase (LMWPTP) during osteoblast differentiation and assessed the modulation of Src kinase by LMWPTP. Methods: Differentiation of MC3T3-E1 pre-osteoblasts was induced by incubation with ascorbic acid and β-glycerophosphate for up to 28 days. Src phosphorylation and LMWPTP expression were analyzed by immunoblotting. Src dephosphorylation in vitro was assessed by incubating immunoprecipitated Src with LMWPTP followed by assay of the residual Src activity using Sam68 as substrate. The importance of LMWPTP in Src dephosphorylation was confirmed by silencing pre-osteoblasts with siRNA-LMWPTP and then assessing Src phosphorylation. Results: Pre-osteoblast differentiation was accompanied by a decrease in phosphorylation of the activator site of Src and an increase in phosphorylation of the inhibitory site. The expression of total Src was unaltered, indicating that post-translational modifications play a pivotal role in Src function. LMWPTP expression was higher in periods when the activator site of Src was dephosphorylated. LMWPTP dephosphorylated pY₅₂₇-Src and pY₄₁₆-Src in vitro, with greater specificity for pY₅₂₇Src. Activation of LMWPTP produced strong activation of Src mediated by fast dephosphorylation of pY₅₂₇-Src, followed by slower deactivation of this kinase via dephosphorylation of pY₄₁₆Src. Conclusion: These results provide new insight into the mechanisms governing the dynamics of Src activity during osteoblast differentiation. A fuller understanding of these mechanisms will improve our knowledge of bone metabolism and of the regulation of Src in other types of cells.

Reduction of Low Molecular Weight Protein-tyrosine Phosphatase Expression Improves Hyperglycemia and Insulin Sensitivity in Obese Mice

To investigate the role of low molecular weight protein-tyrosine phosphatase (LMW-PTP) in glucose metabolism and insulin action, a specific antisense oligonucleotide (ASO) was used to reduce its expression both in vitro and in vivo. Reduction of LMW-PTP expression with the ASO in cultured mouse hepatocytes and in liver and fat tissues of diet-induced obese (DIO) mice and ob/ob mice led to increased phosphorylation and activity of key insulin signaling intermediates, including insulin receptor-beta subunit, phosphatidylinositol 3-kinase, and Akt in response to insulin stimulation. The ASO-treated DIO and ob/ob animals showed improved insulin sensitivity, which was reflected by a lowering of both plasma insulin and glucose levels and improved glucose and insulin tolerance in DIO mice. The treatment did not decrease body weight or increase metabolic rate. These data demonstrate that LMW-PTP is a key negative regulator of insulin action and a potential novel target for the treatment of insulin resistance and type 2 diabetes.

Protein Tyrosine Phosphatase LMW-PTP Exhibits Distinct Roles Between Vascular Endothelial and Smooth Muscle Cells

The present study examined the cellular functions of low-molecular-weight protein tyrosine phosphatase (LMW-PTP), which consists of two active isoforms IF-1 and IF-2, in vascular smooth muscle cells (VSMCs) and endothelial cells (ECs), focusing on cell growth and migration. We transduced recombinant IF-1 and IF-2, and ribozyme targeting both isoforms using an adenovirus vector in these cells. We detected the expression of IF-1 and IF-2 in both types of cells. IF-1 as well as IF-2 inhibited PDGF-induced DNA synthesis and migration in VSMCs. In contrast, both isoforms enhanced lysophosphatidic acid-stimulated cell migration without change in DNA synthesis in ECs. Whereas there is a report indicating that reactive oxygen species-dependent inactivation of LMW-PTP regulates actin cytoskeleton reorganization during cell spreading and migration, the isoforms conversely suppressed the PDGF-induced H2O2 generation with subsequent decrease in the p38 activity in VSMCs. Catalytically inactive LMW-PTP exerted the opposite and similar effects to the wild type in ECs and in VSMCs, respectively, suggesting that substrates for the phosphatase differ between these cells. Moreover, high concentrations of glucose suppressed the expression of LMW-PTP in both cells. These data suggest that LMW-PTP negatively regulates the pathogenesis of atherosclerosis and that glucose-dependent suppression of LMW-PTP expression may promote the development of atherosclerosis in diabetics.

Low Molecular Weight Protein-tyrosine Phosphatase Is Involved in Growth Inhibition during Cell Differentiation

Low molecular weight protein-tyrosine phosphatase (LMW-PTP) is an enzyme involved in mitogenic signaling and cytoskeletal rearrangement after platelet-derived growth factor (PDGF) stimulation. Recently, we demonstrated that LMW-PTP is regulated by a redox mechanism involving the two cysteine residues of the catalytic site, which turn reversibly from reduced to oxidized state after PDGF stimulation. Since recent findings showed a decrease of intracellular reactive oxygen species in contact inhibited cells and a lower tyrosine phosphorylation level in dense cultures in comparison to sparse ones, we studied if the level of endogenous LMW-PTP is regulated by growth inhibition conditions, such as cell confluence and differentiation. Results show that both cell confluence and cell differentiation up-regulate LMW-PTP expression in C2C12 and PC12 cells. We demonstrate that during myogenesis LMW-PTP is regulated at translational level and that the protein accumulates at the plasma membrane. Furthermore, we showed that both myogenesis and cell-cell contact lead to a dramatic decrease of tyrosine phosphorylation level of PDGF receptor. In addition, we observed an increased association of the receptor with LMW-PTP during myogenesis. Herein, we demonstrate that myogenesis decreases the intracellular level of reactive oxygen species, as observed in dense cultures. As a consequence, LMW-PTP turns from oxidized to reduced form during muscle differentiation, increasing its activity in growth inhibition conditions such as differentiation. These data suggest that LMW-PTP plays a crucial role in physiological processes, which require cell growth arrest such as confluence and differentiation.