Changes in the blood level, efficacy, and safety of tacrolimus in pregnancy and the lactation period in patients with systemic lupus erythematosus.

OBJECTIVES: We investigated the efficacy and safety of tacrolimus (TAC) by monitoring its serum concentration for mothers and infants in pregnant patients with systemic lupus erythematosus (SLE). METHODS: We measured trough concentrations of TAC in 25 pregnant patients with SLE to assess influence of TAC on the disease activity. Additionally, we measured the concentrations of TAC in umbilical arterial blood, breast milk, and breastfed infants to investigate the safety of TAC for the mothers and infants. RESULTS: The trough concentrations of TAC in the mothers significantly decreased in the second trimester as compared with those before pregnancy. However, the decrease in the trough concentrations of TAC did not lead to the deterioration of SLE. When examined, the doses of TAC were significantly lower in the second trimester and postpartum in the deteriorating group than those in the non-deteriorating group. There were no adverse events by TAC in mothers and fetuses. The concentrations of TAC in the umbilical cord blood were lower than those in the maternal blood. The relative infant dose in breastfed infants of TAC was < 1%. The level of TAC in infant bloods was below detectable limits. CONCLUSION: These findings suggest that TAC is one of the most effective and safest immunosuppressive drugs for use in pregnant patients with SLE.

Peripartum type B aortic dissection in patients with Marfan syndrome who underwent aortic root replacement: a case series study.

OBJECTIVE: To investigate pregnancy outcomes, especially the risk of pregnancy-related aortic dissection (AD), in patients with Marfan syndrome (MFS) after prophylactic aortic root replacement (ARR). DESIGN: Retrospective case series study. SETTING: Tertiary perinatal care centre at a university hospital. POPULATION: Pregnant women fulfilling the revised Ghent nosology (2010) criteria for MFS who were managed at our institute. METHODS: The pregnancy outcomes of all patients with MFS managed at our institute between 1982 and September 2016 were reviewed retrospectively based on medical records. MAIN OUTCOME MEASURES: Obstetrical management and complication including the incidence of AD throughout the peripartum period. RESULTS: Among 22 patients (28 pregnancies) who had been managed as potential MFS or related disorders, 14 (17 pregnancies) fulfilled the revised Ghent nosology (2010) criteria for MFS and were enrolled in this study. Five patients (five pregnancies) had received ARR before conception: three (60%) developed type B aortic dissection [AD(B)] during the peripartum period, compared with only one of 10 patients (12 pregnancies) without ARR (P < 0.05, Chi-square test). CONCLUSIONS: Our study results suggest that MFS patients after prophylactic ARR are still at high risk of AD(B) during the peripartum period. Careful pre-pregnancy counselling and multidisciplinary care throughout the peripartum period are essential for the management of MFS, even after surgical repair of an ascending aortic aneurysm. TWEETABLE ABSTRACT: MFS patients after prophylactic ARR are still at high risk of type B aortic dissection during the peripartum period.

Influence of antifouling paint on freshwater invertebrates (Mytilidae, Chironomidae and Naididae): density, richness and composition.

We conducted a study about invertebrates on artificial substrates with different antifouling paints in order to answer the following questions 1) is there lower accumulation of organic matter on substrates with antifouling paints, 2) is invertebrate colonization influenced by the release of biocides from antifouling paints, 3) is the colonization of aquatic invertebrates positively influenced by the material accumulated upon the substrate surface and 4) is the assemblage composition of invertebrates similar among the different antifouling paints? To answer these questions, four structures were installed in the Baía River in February 1st, 2007. Each structure was composed of 7 wood boards: 5 boards painted with each type of antifouling paints (T1, T2, T3, T4 and T5), one painted only with the primer (Pr) and the other without any paint (Cn). After 365 days, we observed a greater accumulation of organic matter in the substrates with T2 and T3 paint coatings. Limnoperna fortunei was recorded in all tested paints, with higher densities in the control, primer, T2 and T3. The colonization of Chironomidae and Naididae on the substrate was positively influenced by L. fortunei density. The non-metric multidimensional scaling (NMDS) of the invertebrate community provided evidence of the clear distinction of invertebrate assemblages among the paints. Paints T2 and T3 were the most similar to the control and primer. Our results suggest that antifouling paints applied on substrates hinder invertebrate colonization by decreasing the density and richness of invertebrates.

Influence of antifouling paint on freshwater invertebrates (Mytilidae, Chironomidae and Naididae): density, richness and composition

Abstract We conducted a study about invertebrates on artificial substrates with different antifouling paints in order to answer the following questions 1) is there lower accumulation of organic matter on substrates with antifouling paints, 2) is invertebrate colonization influenced by the release of biocides from antifouling paints, 3) is the colonization of aquatic invertebrates positively influenced by the material accumulated upon the substrate surface and 4) is the assemblage composition of invertebrates similar among the different antifouling paints? To answer these questions, four structures were installed in the Baía River in February 1st, 2007. Each structure was composed of 7 wood boards: 5 boards painted with each type of antifouling paints (T1, T2, T3, T4 and T5), one painted only with the primer (Pr) and the other without any paint (Cn). After 365 days, we observed a greater accumulation of organic matter in the substrates with T2 and T3 paint coatings. Limnoperna fortunei was recorded in all tested paints, with higher densities in the control, primer, T2 and T3. The colonization of Chironomidae and Naididae on the substrate was positively influenced by L. fortunei density. The non-metric multidimensional scaling (NMDS) of the invertebrate community provided evidence of the clear distinction of invertebrate assemblages among the paints. Paints T2 and T3 were the most similar to the control and primer. Our results suggest that antifouling paints applied on substrates hinder invertebrate colonization by decreasing the density and richness of invertebrates.

Resumo O estudo dos invertebrados em substrato artificial com diferentes revestimentos antiincrustantes foi realizado com o intuito de responder as seguintes questões 1) Há menor acumulação de material orgânico nos substratos com revestimentos antiincrustantes? 2) A colonização de invertebrados é afetada pela liberação de biocidas dos revestimentos antiincrustantes? 3) A colonização dos invertebrados aquáticos é facilitada por material acumulado sobre a superfície do substrato? 4) A composição da assembléia é similar entre os diferentes revestimentos? Para realização deste estudo, quatro estruturas foram instaladas em 01 de fevereiro de 2007 no rio Baía. Em cada estrutura foram colocadas sete placas de madeira: cinco placas com aplicação de um tipo de revestimentos antiincrustantes (T1, T2, T3, T4 e T5), uma placa apenas com a aplicação do primer (Pr) e a outra placa permaneceu sem aplicação de nenhum revestimento (Cn). Ao final dos 365 dias em que os substratos ficaram submersos observou-se maior acumulação de material orgânico nos substratos dos revestimentos T2 e T3. Limnoperna fortunei foi registrada em todos os revestimentos testados, com maiores densidades encontradas no controle, primer, T2 e T3. A colonização de Chironomidae e Naididae sobre os substratos foi influenciada pela densidade de L. fortunei. A ordenação (NMDS) evidenciou a separação da assembléia de invertebrados entre os revestimentos. Os revestimentos T2 e T3 foram os mais similares ao controle e ao primer. Os resultados obtidos sugerem que revestimentos antiicrustantes aplicados sobre substratos dificultam a colonização de invertebrados, reduzindo a densidade e riqueza de invertebrados.

Influence of antifouling paint on freshwater invertebrates (Mytilidae, Chironomidae and Naididae): density, richness and composition / Influência de revestimentos antiincrustantes em invertebrados aquáticos (Mytilidae, Chironomidae and Naididae): densidade, riqueza e composição

Abstract We conducted a study about invertebrates on artificial substrates with different antifouling paints in order to answer the following questions 1) is there lower accumulation of organic matter on substrates with antifouling paints, 2) is invertebrate colonization influenced by the release of biocides from antifouling paints, 3) is the colonization of aquatic invertebrates positively influenced by the material accumulated upon the substrate surface and 4) is the assemblage composition of invertebrates similar among the different antifouling paints? To answer these questions, four structures were installed in the Baía River in February 1st, 2007. Each structure was composed of 7 wood boards: 5 boards painted with each type of antifouling paints (T1, T2, T3, T4 and T5), one painted only with the primer (Pr) and the other without any paint (Cn). After 365 days, we observed a greater accumulation of organic matter in the substrates with T2 and T3 paint coatings. Limnoperna fortunei was recorded in all tested paints, with higher densities in the control, primer, T2 and T3. The colonization of Chironomidae and Naididae on the substrate was positively influenced by L. fortunei density. The non-metric multidimensional scaling (NMDS) of the invertebrate community provided evidence of the clear distinction of invertebrate assemblages among the paints. Paints T2 and T3 were the most similar to the control and primer. Our results suggest that antifouling paints applied on substrates hinder invertebrate colonization by decreasing the density and richness of invertebrates.

Resumo O estudo dos invertebrados em substrato artificial com diferentes revestimentos antiincrustantes foi realizado com o intuito de responder as seguintes questões 1) Há menor acumulação de material orgânico nos substratos com revestimentos antiincrustantes? 2) A colonização de invertebrados é afetada pela liberação de biocidas dos revestimentos antiincrustantes? 3) A colonização dos invertebrados aquáticos é facilitada por material acumulado sobre a superfície do substrato? 4) A composição da assembléia é similar entre os diferentes revestimentos? Para realização deste estudo, quatro estruturas foram instaladas em 01 de fevereiro de 2007 no rio Baía. Em cada estrutura foram colocadas sete placas de madeira: cinco placas com aplicação de um tipo de revestimentos antiincrustantes (T1, T2, T3, T4 e T5), uma placa apenas com a aplicação do primer (Pr) e a outra placa permaneceu sem aplicação de nenhum revestimento (Cn). Ao final dos 365 dias em que os substratos ficaram submersos observou-se maior acumulação de material orgânico nos substratos dos revestimentos T2 e T3. Limnoperna fortunei foi registrada em todos os revestimentos testados, com maiores densidades encontradas no controle, primer, T2 e T3. A colonização de Chironomidae e Naididae sobre os substratos foi influenciada pela densidade de L. fortunei. A ordenação (NMDS) evidenciou a separação da assembléia de invertebrados entre os revestimentos. Os revestimentos T2 e T3 foram os mais similares ao controle e ao primer. Os resultados obtidos sugerem que revestimentos antiicrustantes aplicados sobre substratos dificultam a colonização de invertebrados, reduzindo a densidade e riqueza de invertebrados.

Primary prophylaxis to prevent obstetric complications in asymptomatic women with antiphospholipid antibodies: a systematic review.

OBJECTIVE: Obstetric complications are common in patients with antiphospholipid syndrome. However, the impact of antiphosholipid antibodies (aPL) in the pregnancy outcomes of asymptomatic aPL carriers is uncertain. The aim of this systematic review is to assess whether primary prophylaxis is beneficial to prevent obstetric complications during pregnancy in asymptomatic women positive for aPL who have no history of recurrent pregnancy loss or intrauterine fetal death. METHODS: Studies evaluating the effect of prophylactic treatment versus no treatment in asymptomatic pregnant aPL carriers were identified in an electronic database search. Design, population and outcome homogeneity of studies was assessed and meta-analysis was performed. The pooled Mantel-Haenszel relative risk of specific pregnancy outcomes was obtained using random effects models. Heterogeneity was measured with the I(2) statistic. All analyses were conducted using Review Manager 5.3. RESULTS: Data from five studies involving 154 pregnancies were included and three studies were meta-analysed. The risk ratio and 95% confidence interval (CI) of live birth rates, preterm birth, low birth weight and overall pregnancy complications in treated and untreated pregnancies were 1.14 (0.18-7.31); 1.71 (0.32-8.98); 0.98 (0.07-13.54) and 2.15 (0.63-7.33),respectively. Results from the meta-analysis revealed that prophylactic treatment with aspirin is not superior to placebo to prevent pregnancy complications in asymptomatic aPL carriers. CONCLUSION: This systematic review did not find evidence of the superiority of prophylactic treatment with aspirin compared to placebo or usual care to prevent unfavourable obstetric outcomes in otherwise healthy women with aPL during the first pregnancy.

A collision tumour consisting of malignant trichoblastoma and melanosarcoma in a rabbit.

A 7-year-old mixed breed neutered female rabbit (Orytolagus cuniculus) developed a solitary black nodular mass (1 cm in diameter) in the skin of the right flank. Microscopically, the mass consisted of an admixture of neoplastic trichoblasts and melanocytes. The former were arranged as solid, trabecular, island-like and gland-like structures and the cells had oval nuclei with prominent nucleoli and lightly eosinophilic scant cytoplasm. The latter population exhibited prominent nuclear atypia and high mitotic index in the clusters of a few cells or single cells. Immunohistochemically, the neoplastic trichoblasts expressed cytokeratins and E-cadherin, while the neoplastic melanocytes expressed vimentin, S100 protein, melan-A and melanoma antigen. A diagnosis of collision tumour involving malignant trichoblastoma and melanosarcoma was made.

Cardiac hamartoma in a young squirrel monkey who died suddenly.

A case of cardiac hamartoma in a 2-month-old squirrel monkey is reported. The monkey showed a loss of appetite and died suddenly. Microscopically, an encapsulated nodular lesion was found at the right atrial wall. The lesion consisted of irregularly shaped, slender myocytes intermingled with a few fibroblasts and collagen fibers. Neither nuclear atypia nor inflammatory cell infiltrate was seen. The constituting cells had stratified striations in the cytoplasm and reacted immunohistochemically for desmin, indicating the nature of myocytes. Based on the above findings, a diagnosis of cardiac hamartoma was made. This is the first case of cardiac hamartoma in this species.

Developments of scanning probe microscopy with stress/strain fields.

An innovative stress/strain fields scanning probe microscopy in ultra high vacuum (UHV) environments is developed for the first time. This system includes scanning tunneling microscope (STM) and noncontact atomic force microscope (NC-AFM). Two piezo-resistive AFM cantilever probes and STM probes used in this system can move freely in XYZ directions. The nonoptical frequency shift detection of the AFM probe makes the system compact enough to be set in the UHV chambers. The samples can be bent by an anvil driven by a step motor to induce stress and strain on their surface. With a direct current (dc) power source, the sample can be observed at room and high temperatures. A long focus microscope and a monitor are used to observe the samples and the operation of STM and AFM. Silicon(111) surface in room temperature and silicon(001) surface in high temperature with stress were investigated to check the performance of the scanning probe microscope.

Eight years of monitoring aquatic Oligochaeta from the Baía and Ivinhema Rivers.

The aim of this study was to analyze the factors that influence spatial and temporal variations of the Oligochaeta assemblage in the Baía and Ivinhema Rivers, located in the Upper Paraná River floodplain (Mato Grosso do Sul State, Brazil). Samples of Oligochaeta were collected between February 2000 and November 2007. A total of 27 Oligochaeta species were identified. A Principal Component Analysis (PCA) of the physical and chemical variables was used to summarize the total variation in the data and to identify major environmental gradients. Detrended Correspondence Analysis (DCA) was conducted to verify possible spatial and temporal gradients in the Oligochaeta species distribution. The highest Oligochaeta species densities and richness values were recorded during limnophases. The intensity and amplitude of the potamophase influenced the density, richness and composition of Oligochaeta since many species were transported by high water current velocities and/or died due to the low oxygen levels that are characteristic of this phase. L. hoffmeisteri, P. descolei and A. pigueti were recorded in the potamophase and in the limnophase and we concluded that they are adapted to different conditions of rivers (lotic and lentic) due to the presence of these species during the entire study period in both rivers. Nevertheless, these species were dominant in the potamophase periods of these rivers as they have body adaptations to survive in these conditions.

Eight years of monitoring aquatic Oligochaeta from the Baía and Ivinhema Rivers / Oito anos de monitoramento de Oligochaeta aquáticas nos Rios Baía e Ivinhema

The aim of this study was to analyze the factors that influence spatial and temporal variations of the Oligochaeta assemblage in the Baía and Ivinhema Rivers, located in the Upper Paraná River floodplain (Mato Grosso do Sul State, Brazil). Samples of Oligochaeta were collected between February 2000 and November 2007. A total of 27 Oligochaeta species were identified. A Principal Component Analysis (PCA) of the physical and chemical variables was used to summarize the total variation in the data and to identify major environmental gradients. Detrended Correspondence Analysis (DCA) was conducted to verify possible spatial and temporal gradients in the Oligochaeta species distribution. The highest Oligochaeta species densities and richness values were recorded during limnophases. The intensity and amplitude of the potamophase influenced the density, richness and composition of Oligochaeta since many species were transported by high water current velocities and/or died due to the low oxygen levels that are characteristic of this phase. L. hoffmeisteri, P. descolei and A. pigueti were recorded in the potamophase and in the limnophase and we concluded that they are adapted to different conditions of rivers (lotic and lentic) due to the presence of these species during the entire study period in both rivers. Nevertheless, these species were dominant in the potamophase periods of these rivers as they have body adaptations to survive in these conditions.

O objetivo do estudo foi analisar os fatores que influenciam as variações espaciais e temporais da assembléia de Oligochaeta nos Rios Baía e Ivinhema, localizados na planície de inundação do Alto Rio Paraná. (Estado do Mato Grosso do Sul, Brasil). As amostras de Oligochaeta foram coletadas de fevereiro de 2000 a novembro de 2007. Um total de 27 espécies foi identificado. Análise de Componentes Principais (PCA) das variáveis físicas e químicas foi usada para sumarizar a variação total nos dados e identificar os principais gradientes ambientais. Para verificar possíveis gradientes espaciais e temporais na distribuição das espécies de Oligochaeta foi realizada uma Análise de Correspondência Dendrítica com remoção do efeito de arco (DCA). As maiores densidades e riqueza de espécies de Oligochaeta foram registradas nos períodos de águas baixas. A intensidade e amplitude da potamofase influenciaram a riqueza, densidade e composição de espécies, uma vez que muitas das espécies foram carreadas pela alta velocidade de corrente de água e/ou mortas devido aos baixos níveis de oxigênio que foram característicos dessa fase. L. hoffmeisteri, P. descolei and A. pigueti foram registradas nos períodos de potamofase e limnofase e nós concluímos que são adaptadas a diferentes condições de rios (lótico e lêntico) devido a preseça dessas espécies durante todo período estudado em ambos os rios. Contudo, essas espécies foram dominantes nos períodos de potamofase desses rios uma vez que elas apresentam adaptações corporais para sobreviver nessas condições.

Prostaglandin F(2alpha) negatively regulates bone resorption in murine osteoclast development.

Prostaglandin (PG) E(2) promotes osteoclastic cell differentiation, but the physiological function of PGF(2alpha) remains unclear. We examined the physiological effects of PGF(2alpha) on osteoclast differentiation using a murine cell line, RAW264, and the column-purified murine bone marrow cells, both of which are differentiable into osteoclast-like multi-nuclear cells. Although PGF(2alpha) did not affect the number of differentiated osteoclasts, PGF(2alpha) reduced the bone resorption activity of osteoclasts developed from both cell types in a pit formation assay. Thus, PGF(2alpha) inhibits bone resorption without affecting the number of osteoclasts, providing a novel molecular mechanism underlying bone metabolism.

Decreased CHK protein levels are associated with Src activation in colon cancer cells.

Src activation has been associated with colon cancers but the mechanism underlying Src activation is largely unknown. Csk-homologous kinase (CHK) can inhibit the kinase activity of certain Src kinase family members in vitro by phosphorylating the C-terminal tyrosine and by a non-catalytic mechanism. CHK was previously reported to be expressed primarily in brain and hematopoietic cells. We report herein that CHK is also expressed in normal colon cell lines. Furthermore, CHK protein levels are significantly decreased in various colon cancer cell lines and the decrease correlates with the increased specific activity of Src in these cell lines, while the level of the other Src inhibitory kinase, C-terminal Src kinase, is not significantly changed. CHK is also expressed in normal colon tissues but its expression level is decreased in colon cancer tissues collected from the same patients. Immunofluorescence microscopy shows that CHK colocalizes with Src in normal colon FHC cells. Overexpression of CHK in colon cancer cells results in inactivation of Src without phosphorylating Y530 at its C-terminus. In addition, CHK suppresses anchorage-independent cell growth and cell invasion of colon cancer cells. These results reveal a potentially important role for CHK in Src activation and tumorigenicity in colon cancer cells.

An atomic resolution scanning tunneling microscope that applies external tensile stress and strain in an ultrahigh vacuum.

We have developed an ultrahigh vacuum scanning tunneling microscope with an in situ external stress application capability in order to determine the effects of stress and strain on surface atomistic structures. It is necessary to understand these effects because controlling them will be a key technology that will very likely be used in future nanometer-scale fabrication processes. We used our microscope to demonstrate atomic resolution imaging under external tensile stress and strain on the surfaces of wafers of Si(111) and Si(001). We also successfully observed domain redistribution induced by applying uniaxial stress at an elevated temperature on the surface of a wafer of vicinal Si(100). We confirmed that domains for which an applied tensile stress is directed along the dimer bond become less stable and shrink. This suggests that it may be feasible to fabricate single domain surfaces in a process that controls surface stress and strain.

SUMO modification of Sam68 enhances its ability to repress cyclin D1 expression and inhibits its ability to induce apoptosis.

Sam68 (Src associated in mitosis; 68 kDa) is an RNA-binding protein and substrate of Src family kinases. It is thought to play a role in cell cycle progression. Overexpression of Sam68 in fibroblasts was reported to have two separable functions dependent on its ability to bind RNA--cell cycle arrest or the induction of apoptosis. Post-translational modification with SUMO (small ubiquitin-like modifier) is common to many transcription factors and can regulate protein localization, stability and function. Here we show Sam68 to be modified by SUMO, and demonstrate that the SUMO E3 ligase (PIAS1) (protein inhibitor of activated STAT1) can enhance Sam68 sumoylation. Lysine 96, the first lysine in the amino-terminal region of Sam68, was found to be the major SUMO acceptor site. Mutation of the SUMO acceptor lysine to arginine enhanced the ability of Sam68 to induce apoptosis but inhibited its ability to act as a transcriptional inhibitor of cyclin D1 expression. A SUMO-1 Sam68 fusion protein, on the other hand, inhibited the ability of Sam68 to induce apoptosis but was a strong repressor of cyclin D1 expression. Thus, SUMO may be an important regulator of Sam68 function in cell cycle progression.

Prostaglandin E2 induced the differentiation of osteoclasts in mouse osteoblast-depleted bone marrow cells.

Prostaglandin (PG) E(2) is a known bone absorbing agent that acts on osteoblasts to facilitate osteoclastogenesis by increasing the secretion of RANKL. In the present study, we investigated the direct action of PGE(2) on osteoclastic progenitors that differentiate into TRAP-positive multinucleated cells. The hematopoietic stem cell obtained from murine bone marrow was purified by a Sephadex G-10 column, and cultured in the presence of CSF-1 and RANKL to facilitate cell differentiation. The introduction of low-density PGE(2) into the culture resulted in a drastic increase of TRAP-positive multinucleated cells, whereas the addition of high-density PGE(2) had the opposite effect. PCR analysis revealed increased level of EP3 mRNA in undifferentiated cells and reduced level after the development of osteoclast; EP1, EP2 and EP4 were constitutively expressed throughout the differentiation. Investigation of intracellular signaling verified that low-density PGE(2) suppressed PKA activity in undifferentiated cells, suggesting that PGE(2) acts on the osteoclastic cell lineage to facilitate cell differentiation by suppressing PKA in the presence of RANKL.

Selected glimpses into the activation and function of Src kinase.

Since the discovery of the v-src and c-src genes and their products, much progress has been made in the elucidation of the structure, regulation, localization, and function of the Src protein. Src is a non-receptor protein tyrosine kinase that transduces signals that are involved in the control of a variety of cellular processes such as proliferation, differentiation, motility, and adhesion. Src is normally maintained in an inactive state, but can be activated transiently during cellular events such as mitosis, or constitutively by abnormal events such as mutation (i.e. v-Src and some human cancers). Activation of Src occurs as a result of disruption of the negative regulatory processes that normally suppress Src activity, and understanding the various mechanisms behind Src activation has been a target of intense study. Src associates with cellular membranes, in particular the plasma membrane, and endosomal membranes. Studies indicate that the different subcellular localizations of Src could be important for the regulation of specific cellular processes such as mitogenesis, cytoskeletal organization, and/or membrane trafficking. This review will discuss the history behind the discovery and initial characterization of Src and the regulatory mechanisms of Src activation, in particular, regulation by modification of the carboxy-terminal regulatory tyrosine by phosphatases and kinases. Its focus will then turn to the different subcellular localizations of Src and the possible roles of nuclear and perinuclear targets of Src. Finally, a brief section will review some of our present knowledge regarding Src involvement in human cancers.

Identification of protein-tyrosine phosphatase 1B as the major tyrosine phosphatase activity capable of dephosphorylating and activating c-Src in several human breast cancer cell lines.

c-Src tyrosine kinase activity is elevated in several types of human cancer, and this has been attributed to elevated c-Src expression levels, increased c-Src specific activity, and activating mutations in c-Src. We have found a number of human breast cancer cell lines with elevated c-Src specific activity that also possess elevated phosphatase activity directed against the carboxyl-terminal negative regulatory domain of Src family kinases. To identify this phosphatase, cell extracts from MDA-MB-435S cells were chromatographed and the fractions were assayed for phosphatase activity. Four peaks of phosphatase activity directed against the nonspecific substrate poly(Glu/Tyr) were detected. One peak also dephosphorylated a peptide modeled against the c-Src carboxyl-terminal negative regulatory domain and intact human c-Src. Immunoblotting and immunodepletion experiments identified the phosphatase as protein-tyrosine phosphatase 1B (PTP1B). Examination of several human breast cancer cell lines with increased c-Src activity showed elevated levels of PTP1B protein relative to normal control breast cells. In vitro c-Src reactivation experiments confirmed the ability of PTP1B to dephosphorylate and activate c-Src. In vivo overexpression of PTP1B in 293 cells caused a 2-fold increase of endogenous c-Src kinase activity. Our findings indicate that PTP1B is the primary protein-tyrosine phosphatase capable of dephosphorylating c-Src in several human breast cancer cell lines and suggests a regulatory role for PTP1B in the control of c-Src kinase activity.

Human autoantibodies to a novel Golgi protein golgin-67: high similarity with golgin-95/gm 130 autoantigen.

Autoantibodies to subcellular organelles have been described in patients with various systemic rheumatic diseases and our laboratories have been focused on studies of the Golgi complex as the autoimmune target. We have previously isolated and described four of the five known Golgi autoantigens reported to date. During the characterization of Golgi autoantigen golgin-95/gm130, another human cDNA that shared a significant degree of similarity in both nucleotide and amino acid sequences was identified. Analysis of cDNAs from different libraries suggested that this is a distinct gene encoding a protein of 67 kDa which has four regions with sequence identity to gm130, ranging between 42 and 60%. In this report, we describe the complete cDNA encoding a closely related Golgi protein provisionally named golgin-67. Among a group of 84 human anti-Golgi sera, five (6%) were shown to recognize golgin-67. Anti-golgin-67 human sera and affinity purified rabbit antibody to the recombinant protein gave predominant Golgi staining. Golgin-67 is thus the smallest member of a growing family of Golgi autoantigens rich in alpha-helical coiled-coil domain. The current hypothesis for the generation of autoimmune antibody to the Golgi complex is discussed.