Tubulin Post-Translational Modifications: The Elusive Roles of Acetylation.

Microtubules (MTs), dynamic polymers of α/ß-tubulin heterodimers found in all eukaryotes, are involved in cytoplasm spatial organization, intracellular transport, cell polarity, migration and division, and in cilia biology. MTs functional diversity depends on the differential expression of distinct tubulin isotypes and is amplified by a vast number of different post-translational modifications (PTMs). The addition/removal of PTMs to α- or ß-tubulins is catalyzed by specific enzymes and allows combinatory patterns largely enriching the distinct biochemical and biophysical properties of MTs, creating a code read by distinct proteins, including microtubule-associated proteins (MAPs), which allow cellular responses. This review is focused on tubulin-acetylation, whose cellular roles continue to generate debate. We travel through the experimental data pointing to α-tubulin Lys40 acetylation role as being a MT stabilizer and a typical PTM of long lived MTs, to the most recent data, suggesting that Lys40 acetylation enhances MT flexibility and alters the mechanical properties of MTs, preventing MTs from mechanical aging characterized by structural damage. Additionally, we discuss the regulation of tubulin acetyltransferases/desacetylases and their impacts on cell physiology. Finally, we analyze how changes in MT acetylation levels have been found to be a general response to stress and how they are associated with several human pathologies.

Expression of thimet oligopeptidase (THOP) modulated by oxidative stress in human multidrug resistant (MDR) leukemia cells.

Thimet oligopeptidase (THOP) is a cytosolic metallopeptidase known to regulate the fate of post-proteasomal peptides, protein turnover and peptide selection in the antigen presentation machinery (APM) system. Oxidative stress influences THOP expression and regulates its proteolytic activity, generating variable cytosolic peptide levels, possibly affecting the immune evasion of tumor cells. In the present work, we examined the association between THOP expression/activity and stress oxidative resistance in human leukemia cells using the K562 cell line, a chronic myeloid leukemia (CML), and the multidrug-resistant (MDR) Lucena 1 (K562-derived MDR cell line) as model. The Lucena 1 phenotype was validated under vincristine treatment and the relative THOP1 mRNA levels and protein expression compared to K562 cell line. Our data demonstrated increased THOP1 gene and protein levels in K562 cells in contrast to the oxidative-resistant Lucena 1, even after H2O2 treatment, suggesting an oxidative stress dependence in THOP regulation. Further, it was observed higher basal levels of reactive oxygen species (ROS) in K562 compared to Lucena 1 cell line using DHE fluorescent probe. Since THOP activity is dependent on its oligomeric state, we also compared its proteolytic activity under reducing agent treatment, which demonstrated that its function modulation with respect to changes in redox state. Finally, the mRNA expression and FACS analyses demonstrated a reduced expression of MHC I only in K562 cell line. In conclusion, our results highlight THOP redox modulation, which could influence antigen presentation in multidrug resistant leukemia cells.

Colchicine Blocks Tubulin Heterodimer Recycling by Tubulin Cofactors TBCA, TBCB, and TBCE.

Colchicine has been used to treat gout and, more recently, to effectively prevent autoinflammatory diseases and both primary and recurrent episodes of pericarditis. The anti-inflammatory action of colchicine seems to result from irreversible inhibition of tubulin polymerization and microtubule (MT) assembly by binding to the tubulin heterodimer, avoiding the signal transduction required to the activation of the entire NLRP3 inflammasome. Emerging results show that the MT network is a potential regulator of cardiac mechanics. Here, we investigated how colchicine impacts in tubulin folding cofactors TBCA, TBCB, and TBCE activities. We show that TBCA is abundant in mouse heart insoluble protein extracts. Also, a decrease of the TBCA/ß-tubulin complex followed by an increase of free TBCA is observed in human cells treated with colchicine. The presence of free TBCA is not observed in cells treated with other anti-mitotic agents such as nocodazole or cold shock, neither after translation inhibition by cycloheximide. In vitro assays show that colchicine inhibits tubulin heterodimer dissociation by TBCE/TBCB, probably by interfering with interactions of TBCE with tubulin dimers, leading to free TBCA. Manipulation of TBCA levels, either by RNAi or overexpression results in decreased levels of tubulin heterodimers. Together, these data strongly suggest that TBCA is mainly receiving ß-tubulin from the dissociation of pre-existing heterodimers instead of newly synthesized tubulins. The TBCE/TBCB+TBCA system is crucial for controlling the critical concentration of free tubulin heterodimers and MT dynamics in the cells by recycling the tubulin heterodimers. It is conceivable that colchicine affects tubulin heterodimer recycling through the TBCE/TBCB+TBCA system producing the known benefits in the treatment of pericardium inflammation.

MOB: Pivotal Conserved Proteins in Cytokinesis, Cell Architecture and Tissue Homeostasis.

The MOB family proteins are constituted by highly conserved eukaryote kinase signal adaptors that are often essential both for cell and organism survival. Historically, MOB family proteins have been described as kinase activators participating in Hippo and Mitotic Exit Network/ Septation Initiation Network (MEN/SIN) signaling pathways that have central roles in regulating cytokinesis, cell polarity, cell proliferation and cell fate to control organ growth and regeneration. In metazoans, MOB proteins act as central signal adaptors of the core kinase module MST1/2, LATS1/2, and NDR1/2 kinases that phosphorylate the YAP/TAZ transcriptional co-activators, effectors of the Hippo signaling pathway. More recently, MOBs have been shown to also have non-kinase partners and to be involved in cilia biology, indicating that its activity and regulation is more diverse than expected. In this review, we explore the possible ancestral role of MEN/SIN pathways on the built-in nature of a more complex and functionally expanded Hippo pathway, by focusing on the most conserved components of these pathways, the MOB proteins. We discuss the current knowledge of MOBs-regulated signaling, with emphasis on its evolutionary history and role in morphogenesis, cytokinesis, and cell polarity from unicellular to multicellular organisms.

[Hemicorporectomy anesthesia: case report]. / Anestesia para hemicorporectomia: relato de caso.

INTRODUCTION: Hemicorporectomy progresses with hemodynamic and ventilatory repercussions that make anesthesia management definitive to patient outcome. OBJECTIVE: Report anesthesia approach for a patient with squamous cell carcinoma submitted to urgent hemicorporectomy after an episode of hypovolemic shock. CASE REPORT: After lesion bleeding, the patient presented hypovolemic shock class 3, and was submitted to urgent procedure under general inhalation anesthesia and intravenous multimodal analgesia, presenting hemodynamic instability requiring massive blood transfusion after spinal cord transection and removal of surgical specimen. CONCLUSION: Anesthetic management is essential in scenarios such as the one reported to assure patient survival.

Hemicorporectomy anesthesia: case report / Anestesia para hemicorporectomia: relato de caso

Abstract Introduction: Hemicorporectomy progresses with hemodynamic and ventilatory repercussions that make anesthesia management definitive to patient outcome. Objective: Report anesthesia approach for a patient with squamous cell carcinoma submitted to urgent hemicorporectomy after an episode of hypovolemic shock. Case report: After lesion bleeding, the patient presented hypovolemic shock class 3, and was submitted to urgent procedure under general inhalation anesthesia and intravenous multimodal analgesia, presenting hemodynamic instability requiring massive blood transfusion after spinal cord transection and removal of surgical specimen. Conclusion: Anesthetic management is essential in scenarios such as the one reported to assure patient survival.

Resumo Introducão: A hemicorporectomia cursa com repercussões hemodinâmicas e ventilatórias que fazem o manejo anestésico ser definitivo para o desfecho do paciente. Objetivo: Relatar a condução anestésica em um portador de carcinoma espinocelular submetido à hemicorporectomia de urgência após episódio de choque hipovolêmico. Relato de caso: Após sangramento pela lesão, paciente apresentou choque hipovolêmico classe 3, sendo submetido à abordagem de urgência sob anestesia geral inalatória e analgesia multimodal endovenosa, apresentando instabilidade hemodinâmica com necessidade de transfusão sanguínea maciça após secção medular e retirada da peça cirúrgica. Conclusão: O manejo pelo anestesista se faz fundamental em situações como a relatada para assegurar a sobrevida do paciente.

Polarity in Ciliate Models: From Cilia to Cell Architecture.

Tetrahymena and Paramecium are highly differentiated unicellular organisms with elaborated cortical patterns showing a regular arrangement of hundreds to thousands of basal bodies in longitudinal rows that extend from the anterior to the posterior region of the cell. Thus both ciliates exhibit a permanent antero-posterior axis and left-right asymmetry. This cell polarity is reflected in the direction of the structures nucleated around each basal body such as the ciliary rootlets. Studies in these ciliates showed that basal bodies assemble two types of cilia, the cortical cilia and the cilia of the oral apparatus, a complex structure specialized in food capture. These two cilia types display structural differences at their tip domain. Basal bodies possessing distinct compositions creating specialized landmarks are also present. Cilia might be expected to express and transmit polarities throughout signaling pathways given their recognized role in signal transduction. This review will focus on how local polarities in basal bodies/cilia are regulated and transmitted through cell division in order to maintain the global polarity and shape of these cells and locally constrain the interpretation of signals by different cilia. We will also discuss ciliates as excellent biological models to study development and morphogenetic mechanisms and their relationship with cilia diversity and function in metazoans.

Cilia Distal Domain: Diversity in Evolutionarily Conserved Structures.

Eukaryotic cilia are microtubule-based organelles that protrude from the cell surface to fulfill sensory and motility functions. Their basic structure consists of an axoneme templated by a centriole/basal body. Striking differences in ciliary ultra-structures can be found at the ciliary base, the axoneme and the tip, not only throughout the eukaryotic tree of life, but within a single organism. Defects in cilia biogenesis and function are at the origin of human ciliopathies. This structural/functional diversity and its relationship with the etiology of these diseases is poorly understood. Some of the important events in cilia function occur at their distal domain, including cilia assembly/disassembly, IFT (intraflagellar transport) complexes' remodeling, and signal detection/transduction. How axonemal microtubules end at this domain varies with distinct cilia types, originating different tip architectures. Additionally, they show a high degree of dynamic behavior and are able to respond to different stimuli. The existence of microtubule-capping structures (caps) in certain types of cilia contributes to this diversity. It has been proposed that caps play a role in axoneme length control and stabilization, but their roles are still poorly understood. Here, we review the current knowledge on cilia structure diversity with a focus on the cilia distal domain and caps and discuss how they affect cilia structure and function.

Ultrasound dynamics of gastric content volumes after the ingestion of coconut water or a meat sandwich. A randomized controlled crossover study in healthy volunteers / Dinâmica ultrassonográfica dos volumes do conteúdo gástrico após a ingestão de água de coco ou sanduíche de carne. Um estudo cruzado controlado e randômico com voluntários saudáveis

Abstract Background: Adequate preoperative fasting is critical in preventing pulmonary aspiration of gastric content. We proposed to study the sonographic gastric content dynamics after the ingestion of liquid or solid food in healthy volunteers and confront it with current guidelines for preoperative fasting times. Methods: We performed a prospective, crossover, evaluator-blinded study involving 17 healthy volunteers of both sexes. Each participant fasted for 10 h and was subjected to a baseline gastric ultrasound, intake of 400 mL of coconut water or a 145 g, 355 kcal meat sandwich, and sonographic gastric evaluations after 10 min and every hour until the stomach was completely empty. Results: At baseline, all subjects had an empty stomach. At 10 min, gastric content [mean + standard deviation (SD)] was 240.4 + 69.3 and 248.2 + 119.2 mL for liquid and solid foods, respectively (p > 0.05). Mean + SD gastric emptying times were 2.5 + 0.7 and 4.5 + 0.9 h for liquid and solid foods, respectively (p < 0.001). For the drink, the stomach was completely empty in 59% and 100% of the subjects after two and four hours, and for the sandwich, 65% and 100% of the subjects after four and seven hours, respectively. Conclusions: Sonographic gastric dynamics for coconut water and a meat sandwich resulted in complete gastric emptying times higher and lower, respectively, than those suggested by current guidelines for preoperative fasting.

Resumo Justificativa: O jejum pré-operatório adequado é fundamental para prevenir a aspiração pulmonar do conteúdo gástrico. Nossa proposta foi avaliar a dinâmica ultrassonográfica do conteúdo gástrico após a ingestão de alimentos líquidos ou sólidos em voluntários sadios e confrontá-la com as diretrizes atuais para os períodos de jejum no pré-operatório. Métodos: Um estudo prospectivo, cruzado e avaliador-cego foi feito com 17 voluntários saudáveis de ambos os sexos. Cada participante jejuou por 10 horas e foi submetido a uma ultrassonografia gástrica na fase basal, ingestão de 400 mL de água de coco ou 355 g de sanduíche de carne e avaliações gástricas ultrassonográficas foram feitas após 10 minutos e a cada hora até o estômago estar completamente vazio. Resultados: Na fase basal, todos os participantes estavam com o estômago vazio. Aos 10 minutos, o conteúdo gástrico [média + desvio-padrão (DP)] foi de 240,4 + 69,3 e 248,2 + 119,2 mL para alimentos líquidos e sólidos, respectivamente (p > 0,05). Os tempos médios de esvaziamento gástrico + DP foram de 2,5 + 0,7 e 4,5 + 0,9 horas para alimentos líquidos e sólidos, respectivamente (p < 0,001). Para a bebida, o estômago ficou completamente vazio em 59% e 100% dos sujeitos após duas e quatro horas; para o sanduíche, o estômago ficou completamente vazio em 65% e 100% dos sujeitos após quatro e sete horas, respectivamente. Conclusões: A dinâmica ultrassonográfica do volume gástrico para água de coco e sanduíche de carne resultou em tempos totais de esvaziamento gástrico maiores e menores, respectivamente, do que os sugeridos pelas diretrizes atuais para o jejum pré-operatório.

[Ultrasound dynamics of gastric content volumes after the ingestion of coconut water or a meat sandwich. A randomized controlled crossover study in healthy volunteers]. / Dinâmica ultrassonográfica dos volumes do conteúdo gástrico após a ingestão de água de coco ou sanduíche de carne. Um estudo cruzado controlado e randômico com voluntários saudáveis.

BACKGROUND: Adequate preoperative fasting is critical in preventing pulmonary aspiration of gastric content. We proposed to study the sonographic gastric content dynamics after the ingestion of liquid or solid food in healthy volunteers and confront it with current guidelines for preoperative fasting times. METHODS: We performed a prospective, crossover, evaluator-blinded study involving 17 healthy volunteers of both sexes. Each participant fasted for 10h and was subjected to a baseline gastric ultrasound, intake of 400mL of coconut water or a 145g, 355kcal meat sandwich, and sonographic gastric evaluations after 10min and every hour until the stomach was completely empty. RESULTS: At baseline, all subjects had an empty stomach. At 10min, gastric content [mean + standard deviation (SD)] was 240.4 + 69.3 and 248.2 + 119.2mL for liquid and solid foods, respectively (p>0.05). Mean + SD gastric emptying times were 2.5 + 0.7 and 4.5 + 0.9h for liquid and solid foods, respectively (p<0.001). For the drink, the stomach was completely empty in 59% and 100% of the subjects after two and four hours, and for the sandwich, 65% and 100% of the subjects after four and seven hours, respectively. CONCLUSIONS: Sonographic gastric dynamics for coconut water and a meat sandwich resulted in complete gastric emptying times higher and lower, respectively, than those suggested by current guidelines for preoperative fasting.

Cardiac arrest after epidural anesthesia for a esthetic plastic surgery: a case report / Parada cardíaca após peridural para cirurgia plástica estética: relato de caso

Abstract Cardiac arrest during neuraxial anesthesia is a serious adverse event, which may lead to significant neurological damage and death if not treated promptly. The associated mechanisms are neglected respiratory failure, extensive sympathetic block, local anaesthetic toxicity, total spinal block, in addition to the growing awareness of the vagal predominance as a predisposing factor. In the case reported, the patient was 25 years old, ASA I, scheduled for a esthetic lipoplasty. After sedation with midazolam and fentany, epidural anesthesia in interspaces T12-L1 and T2-T3 and catheter insertion into inferior puncture were performed. The patient remained in the supine position for 10 min. Then, she was placed in the prone position, developing asystolic cardiac arrest 20 min after the completion of neuraxial blockade. The medical team immediately placed the patient in the supine position and began cardiopulmonary resuscitation. Spontaneous circulation was achieved after twenty minutes of resuscitation. We discuss in this report the exacerbated vagal response as the main event mechanism. The patient's successful outcome emphasizes the importance of anaesthetic monitoring by anesthesiologists, prompt recognition and treatment of rhythm changes on the electrocardiogram.

Resumo A parada cardíaca durante anestesia neuroaxial é um evento adverso grave, que pode ocasionar sequelas neurológicas importantes e morte se não tratada em tempo hábil. Os mecanismos associados são insuficiência respiratória negligenciada, bloqueio simpático extenso, toxicidade por anestésicos locais, raquianestesia total, além da crescente consciência da predominância vagal como fator predisponente. No caso reportado, a paciente tinha 25 anos e estado físico ASA I e foi programada para lipoplastia estética. Após sedação com midazolam e fentanil, foi feita anestesia peridural nos interespaços T12-L1 e T2-T3 e inserção de cateter na punção inferior. A paciente foi mantida em decúbito dorsal horizontal durante 10 minutos. Em seguida, foi posicionada em decúbito ventral, evoluiu com parada cardíaca em assistolia 20 minutos após o bloqueio do neuroeixo. A equipe médica imediatamente colocou a paciente em decúbito dorsal e iniciou as manobras de ressuscitação cardiorrespiratória. O retorno da circulação espontânea foi obtido após 20 minutos de reanimação. É discutida neste relato a resposta vagal exacerbada como principal mecanismo causal do evento. O sucesso do desfecho da paciente em questão ressalta a importância da vigilância do anestesiologista, e do pronto reconhecimento e tratamento de mudanças de ritmo no eletrocardiograma.

[Cardiac arrest after epidural anesthesia for a esthetic plastic surgery: a case report]. / Parada cardíaca após peridural para cirurgia plástica estética: relato de caso.

Cardiac arrest during neuraxial anesthesia is a serious adverse event, which may lead to significant neurological damage and death if not treated promptly. The associated mechanisms are neglected respiratory failure, extensive sympathetic block, local anaesthetic toxicity, total spinal block, in addition to the growing awareness of the vagal predominance as a predisposing factor. In the case reported, the patient was 25 years old, ASA I, scheduled for aesthetic lipoplasty. After sedation with midazolam and fentany, epidural anesthesia in interspaces T12-L1 and T2-T3 and catheter insertion into inferior puncture were performed. The patient remained in the supine position for 10minutes. Then, she was placed in the prone position, developing asystolic cardiac arrest 20minutes after the completion of neuraxial blockade. The medical team immediately placed the patient in the supine position and began cardiopulmonary resuscitation. Spontaneous circulation was achieved after twenty minutes of resuscitation. We discuss in this report the exacerbated vagal response as the main event mechanism. The patient's successful outcome emphasizes the importance of anaesthetic monitoring by anesthesiologists, prompt recognition and treatment of rhythm changes on the electrocardiogram.

Roles of the Taql and Bsml vitamin D receptor gene polymorphisms in hospital mortality of burn patients.

OBJECTIVE: The aim of this study was to evaluate the roles of the Taql and Bsml vitamin D receptor gene polymorphisms in hospital mortality of burn patients. METHODS: In total, 105 consecutive burn injury patients over 18 years in age who were admitted to the Burn Unit of Bauru State Hospital from January to December 2013 were prospectively evaluated. Upon admission, patient demographic information was recorded and a blood sample was taken for biochemical analysis to identify the presence of the Taql(rs731236) and Bsml(rs1544410) polymorphisms. All of the patients were followed over their hospital stay and mortality was recorded. RESULTS: Eighteen of the patients did not sign the informed consent form, and there were technical problems with genotype analysis for 7 of the patients. Thus, 80 patients (mean age, 42.5±16.1 years) were included in the final analysis. In total, 60% of the patients were male, and 16.3% died during the hospital stay. The genotype frequencies for the Taql polymorphism were 51.25% TT, 41.25% TC and 7.50% CC; for the Bsml polymorphism, they were 51.25% GG, 42.50% GA and 6.25% AA. In logistic regression analysis, after adjustments for age, gender and total body surface burn area, there were no associations between the Taql (OR: 1.575; CI95%: 0.148-16.745; p=0.706) or Bsml (OR: 1.309; CI95%: 0.128-13.430; p=0.821) polymorphisms and mortality for the burn patients. CONCLUSIONS: Our results suggest that the Taql and Bsml vitamin D receptor gene polymorphisms are not associated with hospital mortality of burn patients.

Roles of the Taql and Bsml vitamin D receptor gene polymorphisms in hospital mortality of burn patients

OBJECTIVE: The aim of this study was to evaluate the roles of the Taql and Bsml vitamin D receptor gene polymorphisms in hospital mortality of burn patients. METHODS: In total, 105 consecutive burn injury patients over 18 years in age who were admitted to the Burn Unit of Bauru State Hospital from January to December 2013 were prospectively evaluated. Upon admission, patient demographic information was recorded and a blood sample was taken for biochemical analysis to identify the presence of the Taql(rs731236) and Bsml(rs1544410) polymorphisms. All of the patients were followed over their hospital stay and mortality was recorded. RESULTS: Eighteen of the patients did not sign the informed consent form, and there were technical problems with genotype analysis for 7 of the patients. Thus, 80 patients (mean age, 42.5±16.1 years) were included in the final analysis. In total, 60% of the patients were male, and 16.3% died during the hospital stay. The genotype frequencies for the Taql polymorphism were 51.25% TT, 41.25% TC and 7.50% CC; for the Bsml polymorphism, they were 51.25% GG, 42.50% GA and 6.25% AA. In logistic regression analysis, after adjustments for age, gender and total body surface burn area, there were no associations between the Taql (OR: 1.575; CI95%: 0.148-16.745; p=0.706) or Bsml (OR: 1.309; CI95%: 0.128-13.430; p=0.821) polymorphisms and mortality for the burn patients. CONCLUSIONS: Our results suggest that the Taql and Bsml vitamin D receptor gene polymorphisms are not associated with hospital mortality of burn patients.

Risk of colorectal cancer associated with the C677T polymorphism in 5,10-methylenetetrahydrofolate reductase in Portuguese patients depends on the intake of methyl-donor nutrients.

BACKGROUND: Polymorphisms located in genes involved in the metabolism of folate and some methyl-related nutrients are implicated in colorectal cancer (CRC). OBJECTIVE: We evaluated the association of 3 genetic polymorphisms [C677T MTHFR (methylene tetrahydrofolate reductase), A2756G MTR (methionine synthase), and C1420T SHMT (serine hydroxymethyltransferase)] with the intake of methyl-donor nutrients in CRC risk. DESIGN: Patients with CRC (n = 196) and healthy controls (n = 200) matched for age and sex were evaluated for intake of methyl-donor nutrients and the 3 polymorphisms. RESULTS: Except for folate intake, which was significantly lower in patients (P = 0.02), no differences were observed in the dietary intake of other methyl-donor nutrients between groups. High intake of folate (>406.7 microg/d) was associated with a significantly lower risk of CRC (odds ratio: 0.67; 95% CI: 0.45, 0.99). The A2756G MTR polymorphism was not associated with the risk of developing CRC. In contrast, homozygosity for the C677T MTHFR variant (TT) presented a 3.0-fold increased risk of CRC (95% CI: 1.3, 6.7). Similarly, homozygosity for the C1420T SHMT polymorphism also had a 2.6-fold increased risk (95% CI: 1.1, 5.9) of developing CRC. When interactions between variables were studied, low intake of all methyl-donor nutrients was associated with an increased risk of CRC in homozygous participants for the C677T MTHFR polymorphism, but a statistically significant interaction was only observed for folate (odds ratio: 14.0; 95% CI: 1.8, 108.5). No significant associations were seen for MTR or SHMT polymorphisms. CONCLUSION: These results show an association between the C677T MTHFR variant and different folate intakes on risk of CRC.

5' and 3' UTR thymidylate synthase polymorphisms modulate the risk of colorectal cancer independently of the intake of methyl group donors.

Thymidylate synthase, as a rate-limiting step in DNA synthesis, catalyses the conversion of dUMP into dTMP using 5,10-methylenotetrahydrofolate as the methyl donor. Two polymorphisms have been described in this gene: a repeat polymorphism in the 5' promoter enhancer region (3R versus 2R) and a 6 bp deletion in the 3' unstranslated region. Both of these may affect protein levels. The present case control study was aimed at investigating the influence of these two polymorphisms on the development of colorectal cancer (CRC), as well as their potential interaction with folate, vitamin B6 and vitamin B12 intake. A total of 196 cases and 200 controls, matched for age and sex distribution, were included in the study. No association was found between CRC and the 28 bp repeat polymorphism, but it was observed that individuals with the 6 bp/del and del/del genotypes had a significantly lower risk of developing the disease (OR=0.47; 95% CI 0.30-0.72). A combined genotype (2R/2R; 6 bp/del+del/del) was also found, which was associated with an even lower risk of developing of the disease (OR=0.42; 95% CI 0.26-0.69). No significant interaction between these polymorphisms and vitamin intake was observed. These results indicate for the first time that the 6 bp/del allele might be a protective factor in the development of CRC, independent of the intake of methyl group donors.

Molecular cloning, expression and immunolocalization of a novel human cementum-derived protein (CP-23).

Cementum is a unique mineralized connective tissue that covers the root surfaces of the teeth. The cementum is critical for appropriate maturation of the periodontium, both during development as well as that associated with regeneration of periodontal tissues, IU; however, one major impediment to understand the molecular mechanisms that regulate periodontal regeneration is the lack of cementum markers. Here we report on the identification and characterization of one such differentially human expressed gene, termed "cementum protein-23" (CP-23) that appears to be periodontal ligament and cementum-specific. We screened human cementum tumor-derived cDNA libraries by transient expression in COS-7 cells and "panning" with a rabbit polyclonal antibody against a cementoblastoma conditioned media-derived protein (CP). One isolated cDNA, CP-23, was expressed in E. coli and polyclonal antibodies against the recombinant human CP-23 were produced. Expression of CP-23 protein by cells of the periodontium was examined by Northern blot and in situ hybridization. Expression of CP-23 transcripts in human cementoblastoma-derived cells, periodontal ligament cells, human gingival fibroblasts and alveolar bone-derived cells was determined by RT-PCR. Our results show that we have isolated a 1374-bp human cDNA containing an open reading frame that encodes a polypeptide with 247 amino acid residues, with a predicted molecular mass of 25.9 kDa that represents CP species. The recombinant human CP-23 protein cross-reacted with antibodies against CP and type X collagen. Immunoscreening of human periodontal tissues revealed that CP-23 gene product is localized to the cementoid matrix of cementum and cementoblasts throughout the entire surface of the root, cell subpopulations of the periodontal ligament as well as cells located paravascularly to the blood vessels into the periodontal ligament. Furthermore, 98% of putative cementoblasts and 15% of periodontal ligament cells cultured in vitro expressed CP-23 gene product. Cementoblastoma cells and periodontal ligament cells contained a 5.0 kb CP-23 mRNA. In situ hybridization showed strong expression of CP-23 mRNA on cementoblast, cell subpopulations of the periodontal ligament and cells located around blood vessels into the periodontal ligament. Our results demonstrate that CP-23 represents a novel, tissue-specific-gene product being expressed by periodontal ligament subpopulations and cementoblasts. These findings offer the possibility to determine the cellular and molecular events that regulate the cementogenesis process during root development. Furthermore, it might provide new venues for the design of translational studies aimed at achieving predictable new cementogenesis and regeneration of the periodontal tissues.

Papilomavírus humano - HPV em mulheres com citologia oncológica dentro dos limites da normalidade / Human papilomavirus of women with oncotic cytologic normal

Verifica a prevalência de HPV em mulheres com citologia oncológica dentro da normalidade. A prevalência de HPV não foi muito elevada, talvez porque se tratasse de mulheres na faixa de maior risco, que são as mulheres mais jovens