Academic literature on the topic 'Proto-Oncogene Proteins p21(ras)'

The presence and role of c-ras proteins were investigated in mature human sperm cells. The v-H-ras monoclonal antibody (mAb) against the c-ras protein, p21, reacted specifically with the acrosomal region of methanol-fixed as well as unfixed-live capacitated and non-capacitated human sperm cell in the indirect immunofluorescence technique. The v-H-ras mAb predominantly recognized c-ras protein of 21 kDa on the Western blot of lithium diiodosalicylate (LIS)-solubilized human sperm preparation. The incubation of sperm cells with v-H-ras mAb affected the sperm cell function in the human sperm penetration assay. The antibody significantly reduced the acrosome reaction and release of acrosin activity from the sperm cells. There was no effect of the mAb on percentage motility, although the mAb significantly affected various motility characteristics such as linearity, amplitude of lateral head displacement (ALH) and beat frequency, the motility parameters involved in the hyperactivation phenomenon of sperm cells leading to capacitation and acrosome reaction. These results suggest that the c-ras or c-ras-like proteins are present in mature sperm cell and may have a role in capacitation and/or acrosome reaction of human sperm cell.

We have cloned a DNA fragment from the marine mollusc Aplysia californica, which contains sequences homologous to mammalian ras genes, by screening a genomic library with a viral Ha-ras oncogene probe under conditions of low stringency hybridization. Nucleotide sequencing revealed a putative exon that encodes amino acids sharing 68% homology with residues 5 to 54 of mammalian p21ras polypeptides, and which therefore is likely to encode a ras-like Aplysia protein. The cloned locus, designated Apl-ras, is distinct from the Aplysia rho (ras-homologue) gene and appears to be more closely related to mammalian ras. We used a panel of monoclonal antibodies raised against v-Ha-ras p21 to precipitate an Mr 21,000 protein from extracts of Aplysia nervous tissue, ovotestis, and, to a much lesser degree, buccal muscle. Fluorescence immunocytochemistry revealed that ras-like protein is most abundant in neuronal cell bodies and axon processes, with staining most prominent at plasma membranes. Much less was present in other tissues. The prominence of ras protein in neurons, which are terminally differentiated and non-proliferating, indicates that the control of cell division is not the sole function of this proto-oncogene. The large identified neurons of Aplysia offer the opportunity to examine how ras protein might function in mature nerve cells.

We directly expressed human R-ras 23,000-dalton protein (p23) cDNA in Escherichia coli under the control of the trp promoter. GTP-dependent phosphorylation of a p23 threonine 85 substitution mutant was observed. This result is in direct analogy to the autokinase activity of H-ras and K-ras threonine 59 substitution mutants. Normal p23 protein was detected in the human fibrosarcoma cell line HT1080 by immunoprecipitation with rabbit antibodies raised against an E. coli-expressed R-ras fusion protein. The R-ras p23 protein was found to be 3H labeled in the presence of [9,10(n)-3H]palmitic acid and is associated with the P100 membrane fraction of HT1080 cells. These data suggest that human R-ras p23 has biochemical properties very similar to those of the p21 products of the H-, K-, and N-ras proto-oncogenes. We constructed an R-ras minigene and engineered the expression of normal and mutant alleles from the simian virus 40 early region promoter. Normal and mutant R-ras gene products were authenticated by transient expression in COS-7 cells and immunoprecipitation. The valine 38-substituted R-ras p23 displayed reduced electrophoretic mobility. R-ras p21-like proteins, made by eliminating the first 26 R-ras codons, displayed evident mobility differences between the pro form and mature form, along with a valine 12 substitution-dependent change in electrophoretic mobility. Rat-1 fibroblasts were transfected with normal and mutant R-ras alleles and normal and activated H-ras alleles. Unlike the human T24 bladder oncogene-encoded p21, mutant R-ras alleles do not cause monolayer focus formation or growth in soft agar of rat fibroblasts.

We directly expressed human R-ras 23,000-dalton protein (p23) cDNA in Escherichia coli under the control of the trp promoter. GTP-dependent phosphorylation of a p23 threonine 85 substitution mutant was observed. This result is in direct analogy to the autokinase activity of H-ras and K-ras threonine 59 substitution mutants. Normal p23 protein was detected in the human fibrosarcoma cell line HT1080 by immunoprecipitation with rabbit antibodies raised against an E. coli-expressed R-ras fusion protein. The R-ras p23 protein was found to be 3H labeled in the presence of [9,10(n)-3H]palmitic acid and is associated with the P100 membrane fraction of HT1080 cells. These data suggest that human R-ras p23 has biochemical properties very similar to those of the p21 products of the H-, K-, and N-ras proto-oncogenes. We constructed an R-ras minigene and engineered the expression of normal and mutant alleles from the simian virus 40 early region promoter. Normal and mutant R-ras gene products were authenticated by transient expression in COS-7 cells and immunoprecipitation. The valine 38-substituted R-ras p23 displayed reduced electrophoretic mobility. R-ras p21-like proteins, made by eliminating the first 26 R-ras codons, displayed evident mobility differences between the pro form and mature form, along with a valine 12 substitution-dependent change in electrophoretic mobility. Rat-1 fibroblasts were transfected with normal and mutant R-ras alleles and normal and activated H-ras alleles. Unlike the human T24 bladder oncogene-encoded p21, mutant R-ras alleles do not cause monolayer focus formation or growth in soft agar of rat fibroblasts.

Abstract The p21 RAS product has been implicated as part of the downstream signaling of certain nonreceptor tyrosine kinase oncogenes and several growth factor receptor-ligand interactions. We have reported that the chronic myelogenous leukemia oncogene p210 bcr-abl transforms a growth- factor-dependent myeloid cell line NFS/N1.H7 to interleukin-3 (IL-3) independence. In these p210 bcr-abl-transformed cells (H7 bcr-abl.A54) and in two other murine myeloid cell lines transformed to IL-3 independence by p210 bcr-abl, endogenous p21 RAS is activated as determined by an elevated ratio of associated guanosine triphosphate (GTP)/guanosine diphosphate (GDP), assayed by thin-layer chromatography of the nucleotides eluted from p21 RAS after immunoprecipitation with the Y13–259 antibody. Treatment of p210 bcr-abl-transformed cells with a specific tyrosine kinase inhibitor herbimycin A resulted in diminished tyrosine phosphorylation of p210 bcr-abl and associated proteins, without major reduction in expression of the p210 bcr-abl protein itself. Inhibition of p210 bcr-abl-dependent tyrosine phosphorylation resulted in a reduction of active p21RAS-GTP complexes in the transformed cells, in diminished expression of the nuclear early response genes c-jun and c-fos, and in lower cellular proliferation rate. To further implicate p21 RAS in these functional events downstream of p210 bcr-abl tyrosine phosphorylation, we targeted G- protein function directly by limiting the availability of GTP with the inosine monophosphate dehydrogenase inhibitor, tiazofurin (TR). In p210 bcr-abl-transformed cells treated for 4 hours with TR, in which the levels of GTP were reduced by 50%, but GDP, guanosine monophosphate, and adenosine triphosphate (ATP) were unaffected, p210 bcr-abl tyrosine phosphorylation was at control levels. However, expression of c-fos and c-jun nuclear proto-oncogenes were strongly inhibited and p21 RAS activity was downregulated. These findings show that p210 bcr-abl transduces proliferative signals, in part, through downstream activation of p21 RAS. Furthermore, p21 RAS activity is linked to pathways that regulate c-jun and c-fos expression.

The p21 RAS product has been implicated as part of the downstream signaling of certain nonreceptor tyrosine kinase oncogenes and several growth factor receptor-ligand interactions. We have reported that the chronic myelogenous leukemia oncogene p210 bcr-abl transforms a growth- factor-dependent myeloid cell line NFS/N1.H7 to interleukin-3 (IL-3) independence. In these p210 bcr-abl-transformed cells (H7 bcr-abl.A54) and in two other murine myeloid cell lines transformed to IL-3 independence by p210 bcr-abl, endogenous p21 RAS is activated as determined by an elevated ratio of associated guanosine triphosphate (GTP)/guanosine diphosphate (GDP), assayed by thin-layer chromatography of the nucleotides eluted from p21 RAS after immunoprecipitation with the Y13–259 antibody. Treatment of p210 bcr-abl-transformed cells with a specific tyrosine kinase inhibitor herbimycin A resulted in diminished tyrosine phosphorylation of p210 bcr-abl and associated proteins, without major reduction in expression of the p210 bcr-abl protein itself. Inhibition of p210 bcr-abl-dependent tyrosine phosphorylation resulted in a reduction of active p21RAS-GTP complexes in the transformed cells, in diminished expression of the nuclear early response genes c-jun and c-fos, and in lower cellular proliferation rate. To further implicate p21 RAS in these functional events downstream of p210 bcr-abl tyrosine phosphorylation, we targeted G- protein function directly by limiting the availability of GTP with the inosine monophosphate dehydrogenase inhibitor, tiazofurin (TR). In p210 bcr-abl-transformed cells treated for 4 hours with TR, in which the levels of GTP were reduced by 50%, but GDP, guanosine monophosphate, and adenosine triphosphate (ATP) were unaffected, p210 bcr-abl tyrosine phosphorylation was at control levels. However, expression of c-fos and c-jun nuclear proto-oncogenes were strongly inhibited and p21 RAS activity was downregulated. These findings show that p210 bcr-abl transduces proliferative signals, in part, through downstream activation of p21 RAS. Furthermore, p21 RAS activity is linked to pathways that regulate c-jun and c-fos expression.

Abstract Activated ras proto-oncogenes contribute to the pathogenesis of many animal and human malignancies. ras proto-oncogenes are generally activated by point mutations within codons 12 or 61, which result in the expression of ras protein (p21) bearing characteristic single amino acid substitutions at the corresponding residues. The purpose of the current study was to determine whether the presence of single transforming amino acid substitutions can render normal ras protein immunogenic and, thus, a possible target for T cell-mediated tumor therapy. In initial experiments, C57BL/6 mice were immunized with a synthetic peptide corresponding to residues 5 through 16 of p21 containing the transforming substitution of arginine for normal glycine at residue 12. The results demonstrated that class II MHC-restricted T cells which were specific for the peptide could be elicited, and that the peptide-induced T cells could specifically recognize the corresponding intact p21 ras protein. Recognition of p21 ras protein by peptide-specific T cells implies that C57BL/6 APC can process the activated ras protein in a fashion that allows presentation of digested protein by class II MHC molecules in a configuration similar to the configuration with synthetic peptide. Evaluation of the immunogenicity of peptides containing alternative transforming amino acid substitutions of ras protein demonstrated that some, but not all, were immunogenic in individual strains of mice. Therefore, although ras protein-specific T cells can be elicited by immunization with synthetic peptides, not all of the potential ras mutations commonly associated with malignancy may be recognizable by T cells from all individuals.

The human Ki-ras gene was previously reported to contain two alternative fourth exons which encode two distinct p21 proteins differing only at their carboxy termini. The present study shows that either p21 protein is able on its own to transform NIH 3T3 cells to a tumorigenic state.

The human Ki-ras gene was previously reported to contain two alternative fourth exons which encode two distinct p21 proteins differing only at their carboxy termini. The present study shows that either p21 protein is able on its own to transform NIH 3T3 cells to a tumorigenic state.

Transforming Harvey (Ha) ras oncogene products accelerated the time course of Xenopus oocyte maturation induced by insulin, insulinlike growth factor 1, or progesterone. The transforming constructs, [Val-12]Ha p21 and [Val-12, Thr-59]Ha p21, displayed equal potency and efficacy in their abilities to accelerate the growth peptide-induced response. Normal Ha p21 was only 60% as powerful and one-fifth as potent as the mutants containing valine in the 12 position. In contrast, two nontransforming constructs, [Val-12, Ala-35, Leu-36, Thr-59]Ha p21 and [Val-12, Thr-59]Ha(term-174) p21, had no effect on the time course of hormone-induced maturation. Effects of the transforming ras proteins on hormone-induced maturation correlated with their abilities to stimulate in vivo phosphodiesterase activity measured after microinjection of 200 microM cyclic [3H] AMP. When p21 injection followed 90 min of insulin treatment, there was no increase in phosphodiesterase activity over that measured after hormone treatment or p21 injection alone, but additive effects of p21 and insulin on enzyme activity were observed during the first 90 min of insulin treatment. Even though normal Ha p21 and transforming [Val-12, Thr-59]Ha p21 stimulated oocyte phosphodiesterase to equal levels when coinjected with substrate at the initiation of the in vivo assay, the transforming protein elicited a more sustained stimulation of enzyme activity. These results suggest that stimulation of a cyclic AMP phosphodiesterase activity associated with insulin-induced maturation is involved in the growth-promoting actions of ras oncogene products in Xenopus oocytes.

A incidência do melanoma cutâneo em pacientes adultos jovens tem aumentado consideravelmente nos últimos anos. Há, contudo, carência de conhecimentos clinicopatológicos e moleculares sobre os melanomas que ocorrem nessa faixa etária. O presente estudo teve por objetivo avaliar 132 casos de melanoma cutâneo primário em pacientes com idade entre 18 e 30 anos, com ênfase no estudo das características clínicas, histopatológicas e avaliação molecular das mutações nos genes BRAF, NRAS e KIT. Em relação aos achados clínicos e histopatológicos, houve predomínio de indivíduos do sexo feminino (61,4%), sendo o tronco o sítio anatômico mais comumente envolvido (44,3%) e o melanoma extensivo superficial o tipo histológico predominante (79,5%). A mutação V600E no gene BRAF (BRAFV600E) foi analisada em 93 casos, utilizando-se a técnica de RT-PCR. Essa mutação foi identificada em 38,7% (36/93) e, estatisticamente, associada à fase vertical de crescimento (p = 0,01), infiltrado inflamatório discreto (p = 0,02) e presença de mitose intradérmica (p = 0,004). Houve, ainda, forte indício de associação com a presença de ulceração (p = 0,05). Todas essas variáveis apresentaram associação com pior prognóstico do melanoma cutâneo. Observou-se predomínio da mutação BRAFV600E em regiões anatômicas relacionadas à exposição solar intermitente. Nenhum caso de melanoma com fenômeno de regressão apresentou mutação BRAFV600E (p < 0,05). Não houve associação significativa entre BRAFV600E e sexo, tipo histológico, nível de Clark, índice de Breslow, elastose solar, invasão angiolinfática e perineural, satelitose, nevo melanocítico coexistente e sobrevida. A pesquisa de mutações NRAS, pela técnica de RT-PCR, detectou frequência de 3,95% (3/76). As três mutações encontradas foram do tipo 61K e ocorreram em pacientes do sexo masculino e em região de cabeça e pescoço. As mutações BRAFV600E e NRAS, quando presentes, eram mutuamente exclusivas. A frequência de mutações KIT, analisadas por sequenciamento, foi de 11,1% (3/27). As três mutações identificadas estavam localizadas no éxon 9 (G510, G498S e 489I). Houve concomitância de casos com mutação KIT tanto com NRAS, como com BRAFV600E. Devido ao pequeno número de casos com mutação em KIT e NRAS, não foi possível estabelecer correlações clínicas e histopatológicas com esses genes. Este estudo é o primeiro a descrever as mutações G510D e G498S no gene KIT em melanomas cutâneos. No presente estudo, a mutação BRAFV600E, em melanomas cutâneos de adultos jovens, correlacionou-se com características anatomoclínicas de pior prognóstico em relação aos melanomas selvagens para BRAFV600E
The incidence of cutaneous melanoma in young adults has dramatically increased in recent years. However, there is scarce data about the clinicopathological and molecular characteristics on the melanomas occurring at this age group. The present study aimed to evaluate 132 patients aged between 18 and 30 years with primary cutaneous melanoma with emphasis on the study of clinical, histopathological characteristics and molecular evaluation of mutations in BRAF, NRAS and KIT genes. Regarding the clinical and histopathological findings, the following results were found: female predominance (61.4%), trunk was the most commonly anatomical site involved (44.3%) and superficial spreading melanoma, was the most common histological type (79.5 %). The V600E mutation in BRAF (BRAFV600E) gene was analyzed in 93 cases, using RT-PCR. It was present in 38.7% (36/93) and statistically related to the vertical growth phase (p = 0.01), mild inflammatory infiltration (p = 0.02) and the presence of intradermal mitosis (p = 0.004). There was, also, strongly evidence of an association with the presence of ulceration (p = 0.05). Worse prognosis was associated with these variables. There was a predominance of BRAFV600E mutation in anatomical regions related to intermittent sun exposure. No cases of melanoma with BRAFV600E mutation showed regression phenomenon (p < 0.05). There was no significant association between BRAFV600E and gender, histological type, Clark level, Breslow thickness, solar elastosis, angiolymphatic and perineural invasion, sattelitosis, coexisting melanocytic nevus and survival. The presence of a mutation in NRAS, by RT-PCR was seen in 3.95% (3/76) of the cases. All these three mutations were of type 61K, occurred in male patients and the head and neck region. BRAFV600E and NRAS mutations, when present, were mutually exclusive. The frequency of KIT mutations, analyzed by sequencing, was 11.1% (3/27). The three mutations identified in this gene were located in exon 9 (G510, G498S and 489I). Concomitant mutations were found between KIT and NRAS and BRAFV600E. Due to the small number of KIT and NRAS mutated cases, it was not possible to establish clinical and histopathological correlations and mutation status in these genes. This study was the first to describe the G510D and G498S mutations in KIT gene in cutaneous melanomas. In the present study, BRAFV600E mutation in cutaneous melanoma of young adults correlated with anatomic and clinical features of worse prognosis compared to wild type

Pancreatic ductal adenocarcinoma (PDAC), the most common form of pancreatic cancer, develops through progression of premalignant pancreatic intraepithelial neoplasias (PanINs). In mouse-models, KRAS-activation in acinar cells induced an acinar-to-ductal metaplasia (ADM), and mutation of the Kras oncogene is believed to initiate PanIN formation. ADM is also promoted by pancreatic injury, which cooperates with activated KRAS to stimulate PanIN and PDAC formation from metaplastic ducts. Our lab, and others, have shown that the downstream PI3K/AKT pathway is important for KRAS-mediated proliferation and survival in vitro and in vivo. Prior studies have demonstrated that full activation of AKT requires both PDK1- mediated phosphorylation of AKTT308 and mTOR complex 2 (mTORC2)-mediated phosphorylation of AKTS473. Given the importance of the PI3K/AKT signaling axis, I hypothesized that mTORC2 is required for KRAS-driven pancreatic tumorigenesis and investigated this relationship in mice by combining pancreasspecific expression of an activated KRASG12D molecule with deletion of the essential mTORC2 subunit RICTOR. In the context of activated KRAS, Rictor-null pancreata developed fewer PanIN lesions; these lesions lacked mTORC2 signaling and their proliferation and progression were impaired. Higher levels of nuclear cyclin dependent kinase inhibitors (CDKIs) were maintained in Rictor-null lesions, and nuclear BMI1, a known regulator of the CDKI Cdkn2a, inversely correlated with their expression.Rictor was not required for KRAS-driven ADM following acute pancreatitis, however the inverse correlation between CDKIs and BMI1 was maintained in this system. Treatment of PDX-Cre;KRASG12D/+;Trp53R172H/+ mice with an mTORC1/2 inhibitor delayed tumor formation, and prolonged the survival of mice with late stage PDAC. Knockdown of Rictor in established PDAC cell lines impaired proliferation and anchorage independent growth supporting a role for mTORC2 in fully transformed cells. These data suggest that mTORC2 cooperates with activated KRAS in the initiation and progression of PanIN lesions and is required for the transformation and maintenance of PDAC. My work illustrates phenotypic differences between pancreatic loss of Rictor and PDK1 in the context of KRAS, broadens our understanding of this signaling node and suggests that mTORC2 may potentially be a viable target for PDAC therapies.

Pancreatic ductal adenocarcinoma (PDAC), the most common form of pancreatic cancer, develops through progression of premalignant pancreatic intraepithelial neoplasias (PanINs). In mouse-models, KRAS-activation in acinar cells induced an acinar-to-ductal metaplasia (ADM), and mutation of the Kras oncogene is believed to initiate PanIN formation. ADM is also promoted by pancreatic injury, which cooperates with activated KRAS to stimulate PanIN and PDAC formation from metaplastic ducts. Our lab, and others, have shown that the downstream PI3K/AKT pathway is important for KRAS-mediated proliferation and survival in vitro and in vivo. Prior studies have demonstrated that full activation of AKT requires both PDK1- mediated phosphorylation of AKTT308 and mTOR complex 2 (mTORC2)-mediated phosphorylation of AKTS473. Given the importance of the PI3K/AKT signaling axis, I hypothesized that mTORC2 is required for KRAS-driven pancreatic tumorigenesis and investigated this relationship in mice by combining pancreasspecific expression of an activated KRASG12D molecule with deletion of the essential mTORC2 subunit RICTOR. In the context of activated KRAS, Rictor-null pancreata developed fewer PanIN lesions; these lesions lacked mTORC2 signaling and their proliferation and progression were impaired. Higher levels of nuclear cyclin dependent kinase inhibitors (CDKIs) were maintained in Rictor-null lesions, and nuclear BMI1, a known regulator of the CDKI Cdkn2a, inversely correlated with their expression.Rictor was not required for KRAS-driven ADM following acute pancreatitis, however the inverse correlation between CDKIs and BMI1 was maintained in this system. Treatment of PDX-Cre;KRASG12D/+;Trp53R172H/+ mice with an mTORC1/2 inhibitor delayed tumor formation, and prolonged the survival of mice with late stage PDAC. Knockdown of Rictor in established PDAC cell lines impaired proliferation and anchorage independent growth supporting a role for mTORC2 in fully transformed cells. These data suggest that mTORC2 cooperates with activated KRAS in the initiation and progression of PanIN lesions and is required for the transformation and maintenance of PDAC. My work illustrates phenotypic differences between pancreatic loss of Rictor and PDK1 in the context of KRAS, broadens our understanding of this signaling node and suggests that mTORC2 may potentially be a viable target for PDAC therapies.

A subfamília p21 RAS de pequenas GTPases, incluindo KRAS, HRAS e NRAS, participa de muitas redes de sinalização, incluindo proliferação celular, organização do citoesqueleto e apoptose, e é o alvo mais freqüente de mutações ativadoras em câncer. Mutações germinativas em KRAS e HRAS causam graves anormalidades desenvolvimentais levando às síndromes de Noonan, cárdio-facial-cutânea e Costello, porem mutações ativadoras germinativas em NRAS não foram descritas até hoje. A síndrome autoimune linfoproliferativa (ALPS) é o mais comum defeito genético de apoptose linfocitária, cursando com autoimunidade e acúmulo excessivo de linfócitos, particularmente do tipo T + CD4- CD8-. As mutações causadoras de ALPS descritas até hoje afetam a apoptose mediada por Fas, uma das vias extrínsecas de apoptose. Nós demonstramos aqui que os principais achados clínicos de ALPS, bem como uma predisposição para tumores hematológicos, podem ser causados por uma mutação heterozigota ativadora G13D no oncogene NRAS, sem causar prejuízo na apoptose mediada por Fas. O aumento na quantidade intracelular de NRAS ativo, ligado a GTP, induziu a um aumento da sinalização na via RAF/MEK/ERK, o que suprimiu a expressão da proteína pró-apoptótica BIM, e atenuou a apoptose intrínseca mitocondrial. Desta forma, uma mutação germinativa ativadora em NRAS causou um fenótipo clinico diferente do visto em pacientes com mutações em outros membros da família p21 RAS, cursando com um defeito imunológico seletivo, sem distúrbios generalizados do desenvolvimento
The p21 RAS subfamily of small GTPases, including KRAS, HRAS, and NRAS, regulates cell proliferation, cytoskeletal organization and other signaling networks, and is the most frequent target of activating mutations in cancer. Activating germline mutations of KRAS and HRAS cause severe developmental abnormalities leading to Noonan, cardio-facial-cutaneous and Costello syndrome, but activating germline mutations of NRAS have not been reported. Autoimmune lymphoproliferative syndrome (ALPS) is the most common genetic disease of lymphocyte apoptosis and causes autoimmunity as well as excessive lymphocyte accumulation, particularly of CD4-, CD8- ab T cells. Mutations in ALPS typically affect Fas-mediated apoptosis, but certain ALPS individuals have no such mutations. We show here that the salient features of ALPS as well as a predisposition to hematological malignancies can be caused by a heterozygous germline Gly13Asp activating mutation of the NRAS oncogene that does not impair Fas-mediated apoptosis. The increase in active, GTP-bound NRAS augmented RAF/MEK/ERK signaling which markedly decreased the pro-apoptotic protein BIM and attenuated intrinsic, nonreceptor-mediated mitochondrial apoptosis. Thus, germline activating mutations in NRAS differ from other p21 Ras oncoproteins by causing selective immune abnormalities without general developmental defects

Introdução: A doença renal crônica continua sendo um desafio no campo da pesquisa médica. Atualmente um interesse crescente tem surgido no intuito de avaliar o potencial de células tronco em retardar o avanço de doenças crônicas progressivas. Material e Métodos: Para determinar o efeito dessas células em um modelo de progressão de doença renal crônica foram usadas células linhagem negativa (Lin ) separadas magneticamente e injetadas em ratos submetidos à injúria renal. Ratos singênicos Fischer 344 foram submetidos à nefrectomia 5/6 (Nx) e divididos em 3 grupos: Nx (não tratados); NxSC1 (submetidos à infusão de 2 106 células Lin no 15º dia de pós-operatório); e NxSC3 (submetidos à infusão de 2 106 células Lin no 15º, 30º e 45º dias de pós-operatório). No 60º dia de pós-operatório clearance de inulina, imunohistoquímica e immunoblotting foram realizados. Resultados: Os animais submetidos à nefrectomia apresentaram redução do clearance de inulina (0,33 ± 0,02 ml/min/100g peso corpóreo), proteinúria (12 ± 0,5 mg/24hs) , anemia e hipertensão (145 ± 7,7 mmHg) compatíveis com doença renal crônica. A infusão de células Lin- resultou em atenuação da proteinúria (p<0,05) com relação aos animais não tratados a despeito de não ter havido diferença nos níveis de pressão arterial e aldosterona plasmática. Esses achados foram similares entre os grupos tratados com uma ou com três infusões de células. Adicionalmente a infusão de células resultou em redução do índice de glomeruloesclerose e da área intersticial relativa (p<0,05), menor infiltração do tecido renal por macrófagos e linfócitos e menor proliferação celular. A expressão tecidual do p21 e de VEGF já foi associada à aceleração da progressão da lesão renal crônica. No nosso modelo ambas as proteínas tiveram sua expressão reduzida. A redução da expressão tecidual de eNOS tem sido implicada na progressão da doença renal. Em nosso modelo houve aumento dessa expressão após infusão das células Conclusões: A infusão de células Linatenuou todos os marcadores de injúria renal em um modelo de doença precoce possivelmente através de um mecanismo imunomodulador.
Progressive renal failure continues to be a challenge. The use of bone marrowderived stem cells (SCs) represents a means of meeting that challenge. We used lineage-negative (Lin-) SCs to test the hypothesis that Lin- cell infusion decreases renal injury. Syngeneic Fischer 344 rats were submitted to 5/6 nephrectomy and divided into 3 groups: Nx (untreated); NxSC1 (receiving 2 × 106 Lin- cells on postnephrectomy day 15); and NxSC3 (receiving 2 × 106 Lin- cells on postnephrectomy days 15, 30 and 45). Controls were unoperated/untreated. On postnephrectomy day 60, clearance studies, immunohistochemistry and immunoblotting were performed. Lin- cell infusion effectively reduced postnephrectomy proteinuria, glomerulosclerosis, anemia, renal infiltration of immune cells and monocyte chemoattractant protein-1 protein expression, as well as decreasing the interstitial area. Immunostaining for proliferating cell nuclear antigen showed that, in comparison with controls, Nx rats presented greater cell proliferation, whereas NxSC1 rats and NxSC3 rats presented less cell proliferation than did Nx rats. Protein expression of p21 and VEGF increased after nephrectomy and decreased after Lin- cell infusion. Protein expression of eNOS reduced after nephrectomy and increased after cell infusion. These data suggest that SC treatment ameliorates progressive end-stage renal disease.

O bócio multinodular (BMN) é definido como aumento da glândula tireóide devido a proliferação de tireócitos e caracteriza-se pela heterogeneidade no crescimento e função das células foliculares. É uma patologia comum, com aumento da prevalência em áreas com deficiência de iodo, sendo este o principal fator etiológico do BMN. O BMN é considerado uma neoplasia benigna da tireóide. A patogênese desta disfunção ainda não foi inteiramente elucidada. Nesta revisão serão abordados os mecanismos envolvidos na patogênese e os principais aspectos etiológicos e clínicos do BMN.
Multinodular goiter (MNG) is an enlargement of the thyroid gland and is characterized by heterogeneity in growth and function of thyroid follicular cells. It is a common pathology, with higher prevalence in iodine deficiency areas. Iodine deficiency is the main etiologic factor for MNG. MNG have been considered a true thyroid neoplasm. The pathogenesis of multinodular goiter is not yet clarified. The purpose of this review is to summarize the current knowledge of MNG with respect to the pathology, etiologic and clinical characteristics.

Pancreatic cancer is the 4th leading cause of cancer related death in the United States with a median survival time of less than 6 months. Pancreatic ductal adenocarcinoma (PDAC) accounts for greater than 85% of all pancreatic cancers, and is marked by early and frequent mutation of the KRAS oncogene, with activating KRAS mutations present in over 90% of PDAC. To date, though, targeting activated KRAS for cancer treatment has been very difficult, and targeted therapies are currently being sought for the downstream effectors of activated KRAS. Activation of KRAS stimulates multiple signaling pathways, including the MEK-ERK and PI3K-AKT signaling cascades, but the role of downstream effectors in pancreatic tumor initiation and progression remains unclear. I therefore used primary pancreatic ductal epithelial cells (PDECs), the putative cell of origin for PDAC, to determine the role of specific downstream signaling pathways in KRAS activated pancreatic tumor initiation. As one third of KRAS wild type PDACs harbor activating mutations in BRAF , and KRAS and BRAF mutations appear to be mutually exclusive, I also sought to determine the effect of activated BRAF (BRAF V600E ) expression on PDECs and the signaling requirements downstream of BRAF. I found that both KRAS G12D and BRAF V600E expressing PDECs displayed increased proliferation relative to GFP expressing controls, as well as increased PDEC survival after challenge with apoptotic stimuli. This survival was found to depend on both the MEK-ERK and PI3K-AKT signaling cascades. Surprisingly, I found that this survival is also dependent on the IGF1R, and that activation of PI3K/AKT signaling occurs downstream of MEK/ERK activation, and is dependent on signaling through the IGF1R. Consistent with this, I find increased IGF2 expression in KRAS G12D and BRAF V600E expressing PDECs, and show that ectopic expression of IGF2 rescues survival in PDECs with inhibited MEK, but not PI3K. Finally, I showed that the expression of KRAS G12D or BRAF V600E in PDECs lacking both the Ink4a/Arf and Trp53 tumor suppressors is sufficient for tumor formation following orthotopic transplant of PDECs, and that IGF1R knockdown impairs KRAS and BRAF-induced tumor formation in this model. In addition to these findings within PDECs, I demonstrate that KRAS G12D or BRAF V600E expressing tumor cell lines differ in MEK-ERK and PI3K-AKT signaling from PDECs. In contrast to KRAS G12D or BRAF V600E expressing PDECs, activation of AKT at serine 473 in the KRAS G12D or BRAF V600E expressing tumor cell lines does not lie downstream of MEK, and only the inhibition of PI3K alone or both MEK and the IGF1R simultaneously results in loss of tumor cell line survival. However, inhibition of MEK, PI3K, or the IGF1R in KRAS G12D or BRAF V600E expressing tumor cell lines also resulted in decreased proliferation relative to DMSO treated cells, demonstrating that all three signaling cascades remain important for tumor cell growth and are therefore viable options for pancreatic cancer therapeutics.