Thrombotic obstruction of the central venous catheter in patients undergoing hematopoietic stem cell transplantation

This is an integrative literature review with the aim of summarizing the prevention measures and treatment of thrombotic obstruction of long-term semi-implanted central venous catheters, in patients undergoing hematopoietic stem cell transplantation. The sample consisted of seven studies, being two randomized controlled clinical trials, three cohort studies and two case series. Regarding the prevention measures, one single study demonstrated effectiveness, which was a cohort study on the oral use of warfarin. In relation to the treatment measures, three studies evidenced effectiveness, one highlighted the efficacy of streptokinase or urokinase, one demonstrated the benefit of using low-molecular-weight heparin and the other treated the obstruction with heparin or urokinase. Catheter patency research shows a restricted evolution that does not follow the evolution of transplantations, mainly regarding nursing care.

In search of mechanisms associated with mesenchymal stem cell-based therapies for acute kidney injury

Acute kidney injury (AKI) is classically described as a rapid loss of kidney function. AKI affects more than 15% of all hospital admissions and is associated with elevated mortality rates. Although many advances have occurred, intermittent or continuous renal replacement therapies are still considered the best options for reversing mild and severe AKI syndrome. For this reason, it is essential that innovative and effective therapies, without side effects and complications, be developed to treat AKI and the end-stages of renal disease. Mesenchymal stem cell (MSC) based therapies have numerous advantages in helping to repair inflamed and damaged tissues and are being considered as a new alternative for treating kidney injuries. Numerous experimental models have shown that MSCs can act via differentiation-independent mechanisms to help renal recovery. Essentially, MSCs can secrete a pool of cytokines, growth factors and chemokines, express enzymes, interact via cell-to-cell contacts and release bioagents such as microvesicles to orchestrate renal protection. In this review, we propose seven distinct properties of MSCs which explain how renoprotection may be conferred: 1) anti-inflammatory; 2) pro-angiogenic; 3) stimulation of endogenous progenitor cells; 4) anti-apoptotic; 5) anti-fibrotic; 6) anti-oxidant; and 7) promotion of cellular reprogramming. In this context, these mechanisms, either individually or synergically, could induce renal protection and functional recovery. This review summarises the most important effects and benefits associated with MSC-based therapies in experimental renal disease models and attempts to clarify the mechanisms behind the MSC-related renoprotection. MSCs may prove to be an effective, innovative and affordable treatment for moderate and severe AKI. However, more studies need to be performed to provide a more comprehensive global understanding of MSC-related therapies and to ensure their safety for future clinical applications.

Cell therapy prevents structural, functional and molecular remodeling of remote non-infarcted myocardium

Background/objectives: Therapy using bone marrow (BM) cells has been tested experimentally and clinically due to the potential ability to restore cardiac function by regenerating lost myocytes or increasing the survival of tissues at risk after myocardial infarction (MI). In this study we aimed to evaluate whether BM-derived mononuclear cell (MNC) implantation can positively influence the post-MI structural remodeling, contractility and Ca(2 +)-handling proteins of the remote non-infarcted tissue in rats. Methods and results: After 48 h of MI induction, saline or BM-MNC were injected. Six weeks later, MI scars were slightly smaller and thicker, and cardiac dilatation was just partially prevented by cell therapy. However, the cardiac performance under hemodynamic stress was totally preserved in the BM-MNC treated group if compared to the untreated group, associated with normal contractility of remote myocardium as analyzed in vitro. The impaired post-rest potentiation of contractile force, associated with decreased protein expression of the sarcoplasmic reticulum Ca2 +-ATPase and phosphorylated-phospholamban and overexpression of Na(+)/Ca(2 +) exchanger, were prevented by BM-MNC, indicating preservation of the Ca(2 +) handling. Finally, pathological changes on remodeled remote tissue such as myocyte hypertrophy, interstitial fibrosis and capillary rarefaction were also mitigated by cell therapy. Conclusions: BM-MNC therapy was able to prevent cardiac structural and molecular remodeling after MI, avoiding pathological changes on Ca(2 +)-handling proteins and preserving contractile behavior of the viable myocardium, which could be the major contributor to the improvements of global cardiac performance after cell transplantation despite that scar tissue still exists.

Heparin and penicillamine–hypotaurine–epinephrine (PHE) solution during bovine in vitro fertilization procedures impair the quality of spermatozoa but improve normal oocyte fecundation and early embryonic development

The presence of heparin and a mixture of penicillamine, hypotaurine, and epinephrine (PHE) solution in the in vitro fertilization (IVF) media seem to be a prerequisite when bovine spermatozoa are capacitated in vitro, in order to stimulate sperm motility and acrosome reaction. The present study was designed to determine the effect of the addition of heparin and PHE during IVF on the quality and penetrability of spermatozoa into bovine oocytes and on subsequent embryo development. Sperm quality, evaluated by the integrity of plasma and acrosomal membranes and mitochondrial function, was diminished (P<0.05) in the presence of heparin and PHE. Oocyte penetration and normal pronuclear formation rates, as well as the percentage of zygotes presenting more than two pronuclei, was higher (P<0.05) in the presence of heparin and PHE. No differences were observed in cleavage rates between treatment and control (P>0.05). However, the developmental rate to the blastocyst stage was increased in the presence of heparin and PHE (P>0.05). The quality of embryos that reached the blastocyst stage was evaluated by counting the inner cell mass (ICM) and trophectoderm (TE) cell numbers and total number of cells; the percentage of ICM and TE cells was unaffected (P>0.05) in the presence of heparin and PHE (P<0.05). In conclusion, this study demonstrated that while the supplementation of IVF media with heparin and PHE solution impairs spermatozoa quality, it plays an important role in sperm capacitation, improving pronuclear formation, and early embryonic development

Adipose tissue-derived mesenchymal stem cells increase skin allograft survival and inhibit Th-17 immune response

Adipose tissue-derived mesenchymal stem cells (ADSC) exhibit immunosuppressive capabilities both in vitro and in vivo. Their use for therapy in the transplant field is attractive as they could render the use of immunosuppressive drugs unnecessary. The aim of this study was to investigate the effect of ADSC therapy on prolonging skin allograft survival. Animals that were treated with a single injection of donor allogeneic ADSC one day after transplantation showed an increase in donor skin graft survival by approximately one week. This improvement was associated with preserved histological morphology, an expansion of CD4(+) regulatory T cells (Treg) in draining lymph nodes, as well as heightened IL-10 expression and down-regulated IL-17 expression. In vitro, ADSC inhibit naïve CD4(+) T cell proliferation and constrain Th-1 and Th-17 polarization. In summary, infusion of ADSC one day post-transplantation dramatically increases skin allograft survival by inhibiting the Th-17 pathogenic immune response and enhancing the protective Treg immune response. Finally, these data suggest that ADSC therapy will open new opportunities for promoting drug-free allograft survival in clinical transplantation.

Participação do nicho endosteal na regulação da hemopoese de camundongos submetidos à desnutrição proteica

O nicho endosteal da medula óssea abriga as células-tronco hemopoéticas (CTH) em quiescência/autorrenovação. As CTH podem ser classificadas em dois grupos: células que reconstituem a hemopoese em longo prazo (LT-CTH) e curto prazo (CT-CTH). Investigamos, neste trabalho, os efeitos da desnutrição proteica (DP) no tecido ósseo e a participação do nicho endosteal na sinalização osteoblasto-CTH. Para tanto, utilizamos camundongos submetidos à DP induzida pelo consumo de ração hipoproteica. Os animais desnutridos apresentaram pancitopenia e diminuição nas concentrações de proteínas séricas e albumina. Quantificamos as CTH por citometria de fluxo e verificamos que os desnutridos apresentaram menor porcentagem de LT-CTH, CT-CTH e de progenitores multipotentes (PMP). Avaliamos a expressão das proteínas CD44, CXCR4, Tie-2 e Notch-1 nas LT-CTH. Observamos diminuição da expressão da proteína CD44 nos desnutridos. Isolamos as células LT-CTH por cell sorting e avaliamos a expressão gênica de CD44, CXCR4 e NOTCH-1. Verificamos que os desnutridos apresentaram menor expressão de CD44. Em relação ao ciclo celular, verificamos maior quantidade de LT-CTH nas fases G0/G1. Caracterizamos as alterações do tecido ósseo femoral, in vivo. Observamos diminuição da densidade mineral óssea e da densidade medular nos desnutridos. A desnutrição acarretou diminuição da área média das seções transversais, do perímetro do periósteo e do endósteo na cortical do fêmur dos animais. E na região trabecular, verificou-se diminuição da razão entre volume ósseo e volume da amostra e do número de trabéculas, aumento da distância entre as trabéculas e prevalência de trabéculas ósseas em formato cilíndrico. Avaliamos a expressão de colágeno, osteonectina (ON) e osteocalcina (OC) por imuno-histoquímica, e de osteopontina (OPN) por imunofluorescência no fêmur e verificamos diminuição da marcação para OPN, colágeno tipo I, OC e ON nos desnutridos. Evidenciamos, pela técnica do Picrosírius, desorganização na distribuição das fibras colágenas e presença de fibras tipo III nos fêmures dos desnutridos, além de maior número de osteoclastos evidenciados pela reação da fosfatase ácida tartarato resistente. Os osteoblastos da região femoral foram isolados por depleção imunomagnética, imunofenotipados por citometria de fluxo e cultivados em meio de indução osteogênica. Observamos menor positividade para fosfatase alcalina e vermelho de alizarina nas culturas dos osteoblastos dos desnutridos. Avaliamos, por Western Blotting, a expressão de colágeno tipo I, OPN, osterix, Runx2, RANKL e osteoprotegerina (OPG), e, por PCR em tempo real, a expressão de COL1A2, SP7, CXCL12, ANGPT1, SPP1, JAG2 e CDH2 nos osteoblastos isolados. Verificamos que a desnutrição acarretou diminuição da expressão proteica de osterix e OPG e menor expressão gênica de ANGPT1. Avaliamos a proliferação das células LSK (Lin-Sca1+c-Kit+) utilizando ensaio de CFSE (carboxifluoresceína succinimidil ester). Foi realizada cocultura de células LSK e osteoblastos (MC3T3-E1) na presença e ausência de anti-CD44. Após uma semana, verificamos menor proliferação das LSK dos desnutridos. O bloqueio de CD44 das LSK do grupo controle diminuiu a proliferação destas em três gerações. Entretanto, nos desnutridos, esse bloqueio não afetou a proliferação. Concluímos que a DP promoveu alterações no tecido ósseo e nas CTH. Entretanto, não podemos afirmar que as alterações observadas no sistema hemopoético foram decorrentes de alterações exclusivas do nicho endosteal.

Produção e caracterização de biomateriais acelulares bioativos obtidos a partir da decelularização de placentas

A bioengenharia de tecidos baseia-se no uso de moléculas bioativas, células-tronco e biomateriais para reparação de tecidos e/ou órgãos. Biomateriais podem ser classificados de acordo com sua origem em sintéticos ou biológicos. Biomateriais biológicos podem ser produzidos por decelularização, que visa a remoção de células da matriz extracelular (MEC), a qual deve manter sua integridade química e física. Placentas são órgãos de grande interesse na bioengenharia de tecidos visto que são descartadas após o parto e possuem grande volume de matriz extracelular. Métodos de decelularização podem ser classificados em químicos, físicos e enzimáticos. Todos conhecidamente causam alterações na MEC, sendo que a associação deles é comumente utilizada. Este trabalho comparou diferentes protocolos e estabeleceu um método mais favorável para a decelularização de placentas caninas, visando a produção de um biomaterial para futuras aplicações clínicas. Inicialmente ambas as porções - materna e fetal - das placentas foram submetidas à 10 protocolos, que avaliaram variáveis como concentração e tempo de incubação em detergentes, diferentes gradientes de temperatura e a influência da perfusão versus imersão das soluções, na MEC remanescente. Com base na transparência do tecido e na ausência de núcleo celular em cortes histológicos, dois protocolos foram selecionados (I e II). Além dos critérios já mencionados, ambos os protocolos foram comparados quanto à quantidade de DNA remanescente na MEC decelularizada e à permanência e distribuição de algumas das proteínas da matriz. O detergente SDS foi o mais eficaz na remoção de células, embora não tenha sido suficiente para promover uma decelularização tecidual completa. O congelamento prévio das placentas requereu um maior tempo de incubação posterior das amostras nos distintos detergentes. Ambos métodos de perfusão e imersão foram eficazes na remoção das células, embora grande concentração de proteínas do citoesqueleto tenham permanecido retidas na matriz. As amostras processadas pelo protocolo I (SDS 1%, 5mM EDTA + 50mM TRIS + 0,5% antibiótico, e Triton X-100 1%) apresentaram maior preservação da organização estrutural da MEC quando comparadas àquelas processadas de acordo com o protocolo II (que diferiu do anterior pela utilização de solução contendo 0,05% tripsina ao invés de 50mM TRIS), esse último método entretanto foi o que melhor removeu as células das placentas, conforme observado em lâminas histológicas e demonstrado pela menor concentração de DNA. Tanto as porções materna quanto fetal submetidas à ambos protocolos, mantiveram as proteínas laminina, fibronectina e colágeno tipo I. O colágeno tipo III foi observado somente na porção fetal. Conclui-se que o protocolo II foi o mais eficaz no processo de decelularização de placentas caninas tendo promovido a remoção do conteúdo celular e diminuição da concentração de DNA na MEC remanescente. No entanto é necessário otimizar o tempo de incubação das placentas em soluções enzimáticas visando maior conservação do arranjo da matriz decelularizada. A análise da capacidade da MEC decelularizada por tal método para ser utilizada em bioengenharia de tecidos ainda deve ser avaliada in vitro e in vivo

Análise da imunoexpressão de Oct- 4 e C44 em lesões odontogênicas epiteliais benignas

benign epithelial odontogenic lesions are great clinical importance entities that develop in the jaws from the tissues that form teeth. It has been shown that in benign and malignant tumors, are present in a large number of tumor stem cells, which has great implications in the development of these lesions. Oct-4 and CD44 have been demos as important markers for tumoral stem cells. The objective of this study was to identify epithelial cells expressing stem cell markers by immunohistochemical expression of Oct-4 and CD44 in a series of cases of benign epithelial odontogenic lesions. The sample was comprised of 20 cases of odontogenic keratocyst (OKC), 20 cases of solid/multicystic ameloblastoma and 20 cases of adenomatoid odontogenic tumor (AOT). The expression of Oct-4 and CD44 was evaluated in epithelial lesions using the percentage of positive cells (PP) and the intensity of expression (IE), being realized the sum of these scores, resulting in Total Immunostaining Score (TIS) ranging 0 to 7. The results were submitted to the appropriate statistical test (nonparametric Kruskal-Wallis and Spearman correlation coefficient). All cases were positive for both markers and most showed high expression of both markers. The analysis of Oct-4 expression revealed no statistically significant differences (p = 0.406) among the studied lesions. Regarding the CD44 expression, there was a statistically significant difference between the cases of ameloblastoma and TOA in relation to the CCO, with the latter show more cases in the score 7 (p = 0.034). In the correlation analysis of the immunoreactivity of both markers in the three lesions studied, there was no statistically significant correlation. The results of this study identified the presence of cells with stemness characteristics arranged at various sites in the epithelial component of the studied lesions suggesting their possible role in the histogenesis and differentiation in benign epithelial odontogenic lesions, thus contributing to the development of these lesions.

Análise da imunoexpressão de Oct- 4 e C44 em lesões odontogênicas epiteliais benignas

benign epithelial odontogenic lesions are great clinical importance entities that develop in the jaws from the tissues that form teeth. It has been shown that in benign and malignant tumors, are present in a large number of tumor stem cells, which has great implications in the development of these lesions. Oct-4 and CD44 have been demos as important markers for tumoral stem cells. The objective of this study was to identify epithelial cells expressing stem cell markers by immunohistochemical expression of Oct-4 and CD44 in a series of cases of benign epithelial odontogenic lesions. The sample was comprised of 20 cases of odontogenic keratocyst (OKC), 20 cases of solid/multicystic ameloblastoma and 20 cases of adenomatoid odontogenic tumor (AOT). The expression of Oct-4 and CD44 was evaluated in epithelial lesions using the percentage of positive cells (PP) and the intensity of expression (IE), being realized the sum of these scores, resulting in Total Immunostaining Score (TIS) ranging 0 to 7. The results were submitted to the appropriate statistical test (nonparametric Kruskal-Wallis and Spearman correlation coefficient). All cases were positive for both markers and most showed high expression of both markers. The analysis of Oct-4 expression revealed no statistically significant differences (p = 0.406) among the studied lesions. Regarding the CD44 expression, there was a statistically significant difference between the cases of ameloblastoma and TOA in relation to the CCO, with the latter show more cases in the score 7 (p = 0.034). In the correlation analysis of the immunoreactivity of both markers in the three lesions studied, there was no statistically significant correlation. The results of this study identified the presence of cells with stemness characteristics arranged at various sites in the epithelial component of the studied lesions suggesting their possible role in the histogenesis and differentiation in benign epithelial odontogenic lesions, thus contributing to the development of these lesions.

Análise da imunoexpressão de OCT4 e CD44 em neoplasias de glândulas salivares menores e maiores

Salivary gland neoplasms exhibit a wide variety of biological behavior and a high morphological diversity raises the interest in researching these lesions. The stem cells are the main source for the generation and maintenance of cell diversity, disorders in the regulation of these cells can lead to the production of altered stem cells, termed cancer stem cells capable of generate the tumor. Researches on cancer stem cells and associated proteins have been developed in some oral cancers; however, their role in salivary gland neoplasms is not well established. Thus, the aim of this study was to identify the tumor parenchyma cells exhibiting stem cell characteristics, by evaluating the immunoreactivity of OCT4 and CD44, in a number of cases of salivary gland neoplasms. The sample consisted of 20 pleomorphic adenomas, 20 mucoepidermoid carcinomas and 20 adenoid cystic carcinoma located in minor and major salivary glands. The expression of OCT4 and CD44 was evaluated by the percentage of positive cells (PP) and the intensity of expression (IE), it is realized the sum of the scores, resulting in the total score immunostaining (PIT) ranging 0-7. All studied cases showed positive expression of OCT4 and CD44 and higher values than the control groups. It was observed that for OCT4 luminal cells and non-luminal were immunostained in the case of pleomorphic adenomas and adenoid cystic carcinoma. Already the immunoreactivity of CD44 was particularly evident in the non-luminal cells of these lesions. In mucoepidermoid carcinomas for both markers, there was immunoreactivity in squamous and intermediate cells and absence of staining mucous cells. For both markers, a statistically significant higher immunostaining was verified in neoplasms located in the major salivary glands compared with lesions in the minor salivary (p<0.001). At the total sample and in the group of minor salivary glands, malignant neoplasms exhibited higher immunoreactivity for OCT4 than pleomorphic adenoma. However, there was no statistically significant difference between the lesions and between their classifications histomorphologic. Analyzing the correlation between OCT4 and CD44 immunoexpressions, a statistically significant moderate positive correlation (r = 0.444) was observed. The high expression of OCT4 and CD44 may indicate that these proteins play an important role in identifying cancer stem cells, allowing a prediction of biological behavior of salivary gland neoplasms.

Uso de células mononucleares da medula óssea no tratamento de tendinites induzidas experimentalmente em equinos

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Carotenogênese em células de Haematococcus pluvialis induzidas pelos estresses luminoso e nutricional

O objetivo deste trabalho foi avaliar as respostas das células de Haematococcus pluvialis ao processo de indução à carotenogênese, sob estresse luminoso e nutricional. As células foram aclimatadas durante 15 dias em meio WC, com aeração com ar atmosférico sintético filtrado e fluxo de 100 mL min-1, intensidade luminosa de 50 µmol fótons m-2 s-1, fotoperíodo de 12 horas e temperatura de 23ºC. Foram comparados dois tratamentos: cultivo nas condições descritas, mas com aumento da intensidade luminosa para 350 µmol fótons m-2 s-1 ; e cultivo nas mesmas condições do tratamento anterior, mas com aeração contendo 4% de CO2. Os tratamentos foram conduzidos em triplicata, durante dez dias. Com a adição de CO2 e o incremento da iluminação, observou-se aumento da razão carotenoides/clorofila e da biomassa celular. As células cessaram a divisão no segundo dia de estresse, quando o nitrato se tornou limitante, e aumentaram significativamente seu biovolume. A excreção de carbono orgânico e a concentração de astaxantina aumentam em resposta à adição de CO2. O estresse por intensidade luminosa, aliado à adição de CO2, otimiza a carotenogênese em H. pluvialis e aumenta a produção de astaxantina.

Caracterização das fases de desenvolvimento de células gigantes induzidas por Meloidogyne exigua em raiz de seringueira (Hevea brasiliensis Muell. Arg.)

The developmental phases of giant cells induced by root-knot nematodes (Meloidogyne exigua) in rubber plant (Hevea brasiliensis) root were studied in relation to its number and size evaluated in eight sample dates. The results were subject to cluster analysis and principal component analysis. Sample dates were clearly distinct regarding giant cell development. As a result, the nematode infestation cycle was characterized by the following sequential phases: initial, equilibrium, choice and final.

Implante autólogo de células mononucleares provenientes de medula óssea notratamento de tendinites induzidas experimentalmente em equinos: avaliação a longo prazo - 120 dias

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Carotenogênese em células de Haematococcus pluvialis induzidas pelos estresses luminoso e nutricional

The objective of this work was to evaluate the responses of Haematococcus pluvialis cells to the carotenogenesis induction process, under light and nutrition stress. Cells were acclimated during 15 days in WC medium, with aeration with synthetic, filtered atmospheric air and flow rate of 100 mL min-1, light intensity of 50 µmol photons m-2 s-1, photoperiod of 12 hours, and temperature of 23ºC. The following two treatments were compared: cultivation under the described conditions, but with increase of light intensity up to 350 µmol photons m-2 s-1 ; and cultivation under the same conditions as the previous treatment, but with aeration containing 4% CO2. The treatments were done in triplicate, during ten days. With the addition of CO2 and the increment in lighting, an increase was observed in the carotenoids/chlorophyll ratio and cell biomass. Cells stopped dividing on the second day of stress, when nitrate became limiting, and significantly increased their biovolume. The excretion of organic carbon and the concentration of astaxanthin increase in response to the addition of CO2. Stress by light intensity combined with CO2 addition optimizes carotenogenesis in H. pluvialis and increases astaxanthin production.