Structural and functional properties of the Kunitz-type and C-terminal domains of Amblyomin-X supporting its antitumor activity.

Amblyomin-X is a Kunitz-type FXa inhibitor identified through the transcriptome analysis of the salivary gland from Amblyomma sculptum tick. This protein consists of two domains of equivalent size, triggers apoptosis in different tumor cell lines, and promotes regression of tumor growth, and reduction of metastasis. To study the structural properties and functional roles of the N-terminal (N-ter) and C-terminal (C-ter) domains of Amblyomin-X, we synthesized them by solid-phase peptide synthesis, solved the X-Ray crystallographic structure of the N-ter domain, confirming its Kunitz-type signature, and studied their biological properties. We show here that the C-ter domain is responsible for the uptake of Amblyomin-X by tumor cells and highlight the ability of this domain to deliver intracellular cargo by the strong enhancement of the intracellular detection of molecules with low cellular-uptake efficiency (p15) after their coupling with the C-ter domain. In contrast, the N-ter Kunitz domain of Amblyomin-X is not capable of crossing through the cell membrane but is associated with tumor cell cytotoxicity when it is microinjected into the cells or fused to TAT cell-penetrating peptide. Additionally, we identify the minimum length C-terminal domain named F2C able to enter in the SK-MEL-28 cells and induces dynein chains gene expression modulation, a molecular motor that plays a role in the uptake and intracellular trafficking of Amblyomin-X.

Effect of resistance training combined with ß-glucan ingestion on bone of ovariectomized mice.

OBJECTIVE: This study aimed to evaluate the effects of resistance training (RT) combined with prebiotic ß-glucan (BG) ingestion (Saccharomyces cerevisiae) in ovariectomized (OVX) mice. METHODS: Forty-eight mice were divided into six treatments: SHAM group and five OVX groups treated with saline (SAL), alendronate (ALE) (0.1 mg/kg), BG (62.5 mg/kg), RT (climbing a ladder with load) and the combination of BG + RT. All treatments were performed three times per week for 8 weeks starting 38 days after ovariectomy. RESULTS: OVX effectiveness was confirmed by a reduction in uterus mass (p < 0.05). Femur weight was higher in the SHAM group, followed by the ALE and RT groups with intermediate values and the BG + RT, BG and SAL groups with the lowest values (p < 0.05). The bone mineral density (BMDDEXA) results were higher for the SHAM and RT groups compared to the other groups (p < 0.05). RT provided similar bone mass and resistance to fracture to ALE and SHAM. We found a similar number of osteocytes in the RT and SHAM groups, which was significantly higher than that found in animals receiving BG (p < 0.05). The BG + RT combination increased calcium bone concentration. CONCLUSION: RT promoted benefits on bone health in the OVX model, which were not complemented by the consumption of BG.

Completion of radical hysterectomy does not improve survival of patients with cervical cancer and intraoperatively detected lymph node involvement: ABRAX international retrospective cohort study.

BACKGROUND: The management of cervical cancer patients with intraoperative detection of lymph node involvement remains controversial. Since all these patients are referred for (chemo)radiation after the surgery, the key decision is whether radical hysterectomy should be completed as originally planned, taking into account an additional morbidity associated with extensive surgical dissection prior to adjuvant treatment. The ABRAX study investigated whether completing a radical uterine procedure is associated with an improved oncological outcome of such patients. PATIENTS AND METHODS: We performed retrospective analyses of 515 cervical cancer patients (51 institutions, 19 countries) who were referred for primary curative surgery between 2005 and 2015 (stage IA-IIB, common tumour types) in whom lymph node involvement was detected intraoperatively. Patients were stratified according to whether the planned uterine surgery was completed (COMPL group, N = 361) or abandoned (ABAND group, N = 154) to compare progression-free survival. Definitive chemoradiation was given to 92.9% patients in the ABAND group and adjuvant (chemo)radiation or chemotherapy to 91.4% of patients in the COMPL group. RESULTS: The risks of recurrence (hazard ratio [HR] 1.154, 95% confidence intervals [CI] 0.799-1.666, P = 0.45), pelvic recurrence (HR 0.836, 95% CI 0.458-1.523, P = 0.56), or death (HR 1.064, 95% CI 0.690-1.641, P = 0.78) were not significantly different between the two groups. No subgroup showed a survival benefit from completing radical hysterectomy. Disease-free survival reached 74% (381/515), with a median follow-up of 58 months. Prognostic factors were balanced between the two groups. FIGO stage and number of pelvic lymph nodes involved were significant prognostic factors in the whole study cohort. CONCLUSION: We showed that the completion of radical hysterectomy does not improve survival in patients with intraoperatively detected lymph node involvement, regardless of tumour size or histological type. If lymph node involvement is confirmed intraoperatively, abandoning uterine radical procedure should be considered, and the patient should be referred for definitive chemoradiation. CLINICAL TRIALS IDENTIFIER: NCT04037124.

Nutritional shortage augments cisplatin-effects on murine melanoma cells.

Melanoma incidence increases every year worldwide and is responsible for 80% of skin cancer deaths. Due to its metastatic potential and resistance to almost any treatments such as chemo, radio, immune and targeted-therapy, the patients still have a poor prognosis, especially at metastatic stage. Considering that, it is crucial to find new therapeutic approaches to overcome melanoma resistance. Here we investigated the effect of cisplatin (CDDP), one of the chemotherapeutic agents used for melanoma treatment, in association with nutritional deprivation in murine melanoma cell lines. Cell death and autophagy were evaluated after the treatment with cisplatin, nutritional deprivation and its association using an in vitro model of murine melanocytes malignant transformation to metastatic melanoma. Our results showed that nutritional deprivation augmented cell death induced by cisplatin in melanoma cells, especially at the metastatic subtype, with slight effects on melanocytes. Mechanistic studies revealed that although autophagy was present at high levels in basal conditions in melanoma cells, was not essential for cell death process that involved mitochondrial damage, reactive oxygen species production and possible glycolysis inhibition. In conclusion, nutritional shortage in combination with chemotherapeutic drugs as cisplatin can be a valuable new therapeutic strategy to overcome melanoma resistance.

Avaliação radiográfica e tomográfica de cães submetidos à artroplastia coxofemoral total híbrida / Radiographic and tomographic evaluation of total hybrid hip replacement in dogs

A artroplastia total da articulação coxofemoral é a técnica cirúrgica mais efetiva para o tratamento da displasia coxofemoral canina grave. Na atualidade, o exame radiográfico é o método mais utilizado para se avaliar esse procedimento em cães, porém possui algumas limitações. Já o exame tomográfico por meio de tomografia computadorizada possibilita a aquisição de imagens sem sobreposições e, dessa forma, a determinação do posicionamento dos componentes protéticos com maior precisão. A presente pesquisa teve como objetivo avaliar, por meio de exames radiográficos e tomográficos, a utilização de uma prótese total modular híbrida da articulação coxofemoral desenvolvida e confeccionada no Brasil, aplicada experimentalmente em cães sadios. Foram utilizados seis cães esqueleticamente maduros, previamente submetidos ao procedimento cirúrgico da articulação coxofemoral esquerda. Os cães foram submetidos ao exame tomográfico aos 360 dias e três anos após a cirurgia. Todos os animais apresentaram bom posicionamento dos componentes protéticos e redução articular adequada. Ao exame tomográfico, entretanto, foram detectadas áreas de preenchimento irregular em alguns animais. Adicionalmente, o exame tomográfico permitiu a detecção e monitoração de áreas de reabsorção óssea ao redor do componente acetabular. Assim, o exame tomográfico mostrou-se útil na avaliação das articulações protéticas, com a vantagem de apresentar ótima definição das imagens e permitir quantificar alterações periprotéticas e mensurar relações articulares, fundamentais para o acompanhamento pós-cirúrgico tardio.(AU)

The total hip arthroplasty is the most effective surgical technique for the treatment of severe canine hip dysplasia. Currently, the radiographic examination is the most common method used to assess this procedure in dogs, but has some limitations. CT scan by computed tomography instead enables the acquisition of images without overlap, thereby determining the positioning of prosthetic components with greater accuracy. This research aimed to evaluate by radiography and CT scan examinations the use of a hybrid modular total hip prosthesis developed and made in Brazil, applied experimentally in healthy dogs. Six skeletally mature dogs, which previously underwent surgery on the left hip joint were used. The dogs underwent CT scan at 360 days and three years after surgery. All animals showed good positioning of prosthetic components, and proper joint reduction. In the CT scan, however, it was possible to detect irregular fill areas in some animals. In addition, CT scan allowed the detection and monitoring of areas of bone resorption around the acetabular component. Therefore, the CT scan was useful in the evaluation of prosthetic joints, with the advantage of optimum sharpness of the images and allowing for the possibility of quantifying peri-prosthetic changes and measure joint, fundamental relations for late postoperative follow-up.(AU)

Apoptosis induced by Aß25-35 peptide is Ca(2+) -IP3 signaling-dependent in murine astrocytes.

Although the accumulation of the neurotoxic peptide ß-amyloid (Aß) in the central nervous system is a hallmark of Alzheimer's disease, whether Aß acts in astrocytes is unclear, and downstream functional consequences have yet to be defined. Here, we show that cytosolic Ca(2+) dysregulation, induced by a neurotoxic fragment (Aß25-35), caused apoptosis in a concentration-dependent manner, leading to cytoplasmic Ca(2+) mobilization from extra- and intracellular sources, mainly from the endoplasmic reticulum (ER) via IP3 receptor activation. This mechanism was related to Aß-mediated apoptosis by the intrinsic pathway because the expression of pro-apoptotic Bax was accompanied by its translocation in cells transfected with GFP-Bax. Aß-mediated apoptosis was reduced by BAPTA-AM, a fast Ca(2+) chelator, indicating that an increase in intracellular Ca(2+) was involved in cell death. Interestingly, the Bax translocation was dependent on Ca(2+) mobilization from IP3 receptors because pre-incubation with xestospongin C, a selective IP3 receptor inhibitor, abolished this response. Taken together, these results provide evidence that Aß dysregulation of Ca(2+) homeostasis induces ER depletion of Ca(2+) stores and leads to apoptosis; this mechanism plays a significant role in Aß apoptotic cell death and might be a new target for neurodegeneration treatments.

Interplay between apoptosis and autophagy, a challenging puzzle: new perspectives on antitumor chemotherapies.

Autophagy is a mechanism of protection against various forms of human diseases, such as cancer, in which autophagy seems to have an extremely complex role. In cancer, there is evidence that autophagy may be oncogenic in some contexts, whereas in others it clearly contributes to tumor suppression. In addition, studies have demonstrated the existence of a complex relationship between autophagy and cell death, determining whether a cell will live or die in response to anticancer therapies. Nevertheless, we still need to complete the autophagy-apoptosis puzzle in the tumor context to better address appropriate chemotherapy protocols with autophagy modulators. Generally, tumor cell resistance to anticancer induced-apoptosis can be overcome by autophagy inhibition. However, when an extensive autophagic stimulus is activated, autophagic cell death is observed. In this review, we discuss some details of autophagy and its relationship with tumor progression or suppression, as well as role of autophagy-apoptosis in cancer treatments.

The role of calcium stores in apoptosis and autophagy.

The mechanisms that regulate programmed cell death, such as apoptosis, and the cellular "self-eating" phenomenon of autophagy, share many regulatory systems and common pathways. These mechanisms have been extensively investigated over the last few years. Some intracellular structures may determine and control the autophagic fate of the cell such as the endoplasmic reticulum, mitochondria, and lysosomes. The coordination and interrelation of these organelles are crucial in maintaining calcium levels and general cellular homeostasis, as well as in regulating cell life and death under physiological and pathological conditions, including cancer, neurodegeneration, and aging. In this review, we discuss the crosstalk between the aforementioned organelles and their influence in apoptotic and autophagic processes.

Avaliação da utilização de medicamentos na prática clínica em um hospital público / Assessment of drugs prescribed to medical in-patients in a public hospital

Através desta pesquisa, objetivou-se estudar o consumo dos medicamentos em pacientes hospitalizados na clínica médica de um hospital público na cidade de Campina Grande (PB). O estudo caracterizou-se como descritivo e exploratório, de caráter transversal, com abordagens quali-quantitativas, e foi constituído por uma amostra de 107 pacientes que iniciaram internação na clínica médica no período de agosto de 2007 a julho de 2008. Os 107 pacientes apresentavam 270 diagnósticos ativos, sendo as doenças do aparelho circulatório as de maior ocorrência. Dos pacientes, 65,4% eram idosos e 3,33% dos medicamentos prescritos foram considerados impróprios para eles. Os pacientes que apresentaram possíveis 107 RAMs totalizaram 43%, com média de 2,32 por paciente; as que afetaram o sistema gastrintestinal dos pacientes foram as identificadas com maior frequência. Houve 42 interações distintas, envolvendo 26 tipos de fármacos. Dessa forma, os resultados podem ser úteis no estímulo ao desenvolvimento de mecanismos de avaliação de processos que visem reduzir esses riscos, aumentando a chance de resultados terapêuticos positivos e benefícios para os pacientes.

We studied the consumption of drugs by in-patients in the medical ward of a public hospital in the city of Campina Grande, PB, Brazil. This paper describes a descriptive / exploratory cross-sectional quali-quantitative study of a sample of 107 patients who were admitted to the general medical ward, from August 2007 to July 2008. The 107 patients were diagnosed with 270 active complaints, mainly diseases of the circulatory system. Most of the patients (65.4%) were elderly and 3.33% of drugs prescribed for them are considered unfit for use in the elderly. Many patients (43%) presented 107 possible Adverse Drug Reactions, with an average of 2.32 per patient, those affecting the gastrointestinal system of the patients being identified most frequently. There were 42 different drug interactions, involving 26 types of drug. We hope these results may be useful in stimulating the development of means to assess drug treatment in hospital, so as to reduce these risks and increase the chance of positive outcomes and therapeutic benefits for the patients.