Implications for Clinical Practice from a Multicenter Survey of Heart Failure Management Centers.

OBJECTIVES: This observational, cross-sectional study based aimed to test whether heart failure (HF)-disease management program (DMP) components are influencing care and clinical decision-making in Brazil. METHODS: The survey respondents were cardiologists recommended by experts in the field and invited to participate in the survey via printed form or email. The survey consisted of 29 questions addressing site demographics, public versus private infrastructure, HF baseline data of patients, clinical management of HF, performance indicators, and perceptions about HF treatment. RESULTS: Data were obtained from 98 centers (58% public and 42% private practice) distributed across Brazil. Public HF-DMPs compared to private HF-DMP were associated with a higher percentage of HF-DMP-dedicated services (79% vs 24%; OR: 12, 95% CI: 94-34), multidisciplinary HF (MHF)-DMP [84% vs 65%; OR: 3; 95% CI: 1-8), HF educational programs (49% vs 18%; OR: 4; 95% CI: 1-2), written instructions before hospital discharge (83% vs 76%; OR: 1; 95% CI: 0-5), rehabilitation (69% vs 39%; OR: 3; 95% CI: 1-9), monitoring (44% vs 29%; OR: 2; 95% CI: 1-5), guideline-directed medical therapy-HF use (94% vs 85%; OR: 3; 95% CI: 0-15), and less B-type natriuretic peptide (BNP) dosage (73% vs 88%; OR: 3; 95% CI: 1-9), and key performance indicators (37% vs 60%; OR: 3; 95% CI: 1-7). In comparison to non- MHF-DMP, MHF-DMP was associated with more educational initiatives (42% vs 6%; OR: 12; 95% CI: 1-97), written instructions (83% vs 68%; OR: 2: 95% CI: 1-7), rehabilitation (69% vs 17%; OR: 11; 95% CI: 3-44), monitoring (47% vs 6%; OR: 14; 95% CI: 2-115), GDMT-HF (92% vs 83%; OR: 3; 95% CI: 0-15). In addition, there were less use of BNP as a biomarker (70% vs 84%; OR: 2; 95% CI: 1-8) and key performance indicators (35% vs 51%; OR: 2; 95% CI: 91,6) in the non-MHF group. Physicians considered changing or introducing new medications mostly when patients were hospitalized or when observing worsening disease and/or symptoms. Adherence to drug treatment and non-drug treatment factors were the greatest medical problems associated with HF treatment. CONCLUSION: HF-DMPs are highly heterogeneous. New strategies for HF care should consider the present study highlights and clinical decision-making processes to improve HF patient care.

Implications for clinical practice from a multicenter survey of heart failure management centers

OBJECTIVES: This observational, cross-sectional study based aimed to test whether heart failure (HF)-disease management program (DMP) components are influencing care and clinical decision-making in Brazil. METHODS: The survey respondents were cardiologists recommended by experts in the field and invited to participate in the survey via printed form or email. The survey consisted of 29 questions addressing site demographics, public versus private infrastructure, HF baseline data of patients, clinical management of HF, performance indicators, and perceptions about HF treatment. RESULTS: Data were obtained from 98 centers (58% public and 42% private practice) distributed across Brazil. Public HF-DMPs compared to private HF-DMP were associated with a higher percentage of HF-DMP-dedicated services (79% vs 24%; OR: 12, 95% CI: 94-34), multidisciplinary HF (MHF)-DMP [84% vs 65%; OR: 3; 95% CI: 1-8), HF educational programs (49% vs 18%; OR: 4; 95% CI: 1-2), written instructions before hospital discharge (83% vs 76%; OR: 1; 95% CI: 0-5), rehabilitation (69% vs 39%; OR: 3; 95% CI: 1-9), monitoring (44% vs 29%; OR: 2; 95% CI: 1-5), guideline-directed medical therapy-HF use (94% vs 85%; OR: 3; 95% CI: 0-15), and less B-type natriuretic peptide (BNP) dosage (73% vs 88%; OR: 3; 95% CI: 1-9), and key performance indicators (37% vs 60%; OR: 3; 95% CI: 1-7). In comparison to non- MHF-DMP, MHF-DMP was associated with more educational initiatives (42% vs 6%; OR: 12; 95% CI: 1-97), written instructions (83% vs 68%; OR: 2: 95% CI: 1-7), rehabilitation (69% vs 17%; OR: 11; 95% CI: 3-44), monitoring (47% vs 6%; OR: 14; 95% CI: 2-115), GDMT-HF (92% vs 83%; OR: 3; 95% CI: 0-15). In addition, there were less use of BNP as a biomarker (70% vs 84%; OR: 2; 95% CI: 1-8) and key performance indicators (35% vs 51%; OR: 2; 95% CI: 91,6) in the non-MHF group. Physicians considered changing or introducing new medications mostly when patients were hospitalized or when observing worsening disease and/or symptoms. Adherence to drug treatment and non-drug treatment factors were the greatest medical problems associated with HF treatment. CONCLUSION: HF-DMPs are highly heterogeneous. New strategies for HF care should consider the present study highlights and clinical decision-making processes to improve HF patient care.

Cell-free therapy with the secretome of adipose tissue-derived stem cells in rats' frozen-thawed ovarian grafts.

The use of secretome may be a new strand of cell therapy, which is equal to or even superior to the injection of live cells, called cell-free therapy. In ovarian transplantation, this approach may be a therapeutic possibility for the ovarian graft in hypoxia. We designed the present study to evaluate whether the cell-free therapy with the secretome of adipose tissue-derived stem cells (ASCs) in rat frozen-thawed ovarian grafts could protect a graft against ischemic injury. A single dose of rat ASCs secretome or vehicle was injected into the bilateral frozen-thawed ovaries of 18 adult female rats immediately after an autologous transplant. Nine animals were used to control the cryopreservation protocol and were evaluated before and after the cryopreservation process. Daily vaginal smears were performed for estrous cycle evaluation until euthanasia on postoperative day 30. Follicle viability by trypan blue, graft morphology by HE, and apoptosis by TUNEL and cleaved-caspase-3 were assessed. No differences were found with respect to estrous cycle resumption and follicle viability (p > 0.05). However, compared with the vehicle-treated grafts, the morphology of the secretome-treated grafts was impaired, showing reduced follicular population and increased apoptosis (p < 0.05). ASC secretome impaired the rat frozen-thawed ovarian graft from ischemic injury. However, more studies are needed to evaluate the factors involved and the possibility of applying the secretome in scaffolds to optimize its use.

Lamellar keratoplasty in rabbits using an allogeneic free omental graft and omentum-derived mesenchymal cells associated with the canine amniotic membrane / Ceratoplastia lamelar em coelhos usando enxerto de omento alógeno livre e células mesenquimais omentais associadas com membrana amniótica canina

ABSTRACT: The objective of this research was to evaluate the clinical and microscopic effects in rabbits of lamellar keratoplasty using allogeneic omentum associated with canine amniotic membrane (AM). Rabbits were divided into two groups: one received the allogeneic free omental graft covered with the AM (OM-graft group), while the other received the AM graft containing omental mesenchymal cells (OM-cell group). Clinical signs were evaluated on different postoperative days. After the clinical assessments, the rabbits were euthanized and their corneas were obtained for histopathology and immunohistochemistry (Ki-67, marker for proliferation). Both groups showed chemosis, blepharospasm, eye discharge, hyperemia, and corneal opacity/edema. Neovascularization was observed in the OM-cell group. Histopathological evaluation revealed epithelial islands within the stroma of OM-cell samples. Thirty days after surgery, complete corneal re-epithelialization had occurred in both groups. The OM-cell group showed more Ki-67 positive cells. The free omentum and its cells, combined with the AM, contributed to corneal repair, a process that was completed 30 days after lamellar keratoplasty.

RESUMO: Objetivou-se, com a pesquisa, avaliar os efeitos clínicos e microscópicos da associação do omento de coelho com a membrana amniótica (AM) canina, na ceratoplastia lamelar em coelhos. Dois grupos foram constituídos: um recebeu enxerto de omento alógeno livre, recoberto por AM (grupo OM- graft); o outro recebeu enxerto de AM contendo células mesenquimais derivadas do omento (grupo OM-cell). Manifestações clínicas foram avaliadas em diferentes tempos de pós-operatórios. Após as avaliações clínicas, coelhos foram submetidos à eutanásia e córneas foram colhidas para histopatologia e imunohistoquímica (Ki-67, marcador de proliferação). Relativamente às manifestações clínicas, ambos os grupos apresentaram sinais de quemose, blefarospasmo, secreção ocular, hiperemia e opacidade/edema. Neovascularização foi observada no grupo OM-cell. Avaliações à histopatologia mostraram que uma amostra de OM-cell apresentou ilhas de epitélio dentro do estroma. Aos 30 dias de pós-operatório, observou-se reepitelização corneal completa, em OM-graft e OM-cell. O grupo OM-cell apresentou mais células positivas para Ki-67. O omento livre e suas células, associados à AM, contribuíram para a reparação corneal, que se completou após 30 dias de ceratoplastia lamelar.

Scaffold-based delivery of adipose tissue-derived stem cells in rat frozen-thawed ovarian autografts: preliminary studies in a rat model.

PURPOSE: This study aimed to evaluate whether a gelatin-based Gelfoam sponge is feasible as a scaffold for adipose tissue-derived stem cell (ASC) therapy in rat frozen-thawed ovarian autografts. METHODS: Two sets of studies were performed. The in vitro set evaluated ASCs' viability in the Gelfoam scaffold at different times of co-culturing (after 24, 48, 72, 96, and 120 h). The in vivo set used 20 12-week-old adult female Wistar rats. Frozen-thawed ovarian grafts were treated with ASCs delivered in Gelfoam scaffolds immediately after an autologous retroperitoneal transplant (ASCs-GS, n = 10). The controls received Gelfoam with a culture medium (GS, n = 10). Assessment of graft quality was conducted by vaginal smears (until euthanasia on the 30th postoperative day), histological analyses, follicular density, and viability and fibrosis. Immunohistochemical staining for VEGF-A expression, vascular network (vWF), apoptosis (caspase-3 and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL)), cell proliferation (Ki-67), and hormone receptors (estrogen and progesterone) were performed. RESULTS: The cells remained viable in Gelfoam for up to 120 h of co-culturing. The graft morphology was similar among the groups. ASC therapy promoted the earlier resumption of the estrous phase (GS 16.6 ± 3 vs. ASCs-GS 12.8 ± 1.3 days) and enhanced estrogen receptors compared with the controls (p < 0.05) without interfering with the quantity and viability of the ovarian follicles, fibrosis, endothelial cells, VEGF immunoexpression, apoptosis, or cell proliferation (p > 0.05). CONCLUSION: The Gelfoam scaffold could be a feasible and safe non-invasive technique for ASC delivery in the treatment of frozen-thawed ovarian autografts. Future studies should evaluate the real benefit of this treatment on the survival and endocrine activity of the graft.

Adipose tissue-derived stem cell therapy in rat cryopreserved ovarian grafts.

The preliminary results of ovarian transplantation in clinical practice are encouraging. However, the follicular depletion caused by ischemic injury is a main concern and is directly related to short-term graft survival. Cell therapy with adipose tissue-derived stem cells (ASCs) could be an alternative to induce early angiogenesis in the graft. This study aimed to evaluate ASCs therapy in rat cryopreserved ovarian grafts. A single dose of rat ASC (rASCs) or vehicle was injected into the bilateral cryopreserved ovaries of twelve adult female rats immediately after an autologous transplant. Daily vaginal smears were performed for estrous cycle evaluation until euthanasia on postoperative day 30. Follicle viability, graft morphology and apoptosis were assessed. No differences were found with respect to estrous cycle resumption and follicle viability (P>0.05). However, compared with the vehicle-treated grafts, the morphology of the ASCs-treated grafts was impaired, with diffuse atrophy and increased apoptosis (P<0.05). ASCs direct injected in the stroma of rat cryopreserved ovarian grafts impaired its morphology although may not interfere with the functional resumption on short-term. Further investigations are necessary to evaluated whether it could compromise their viability in the long-term.

Females transplanted with ovaries subjected to hypoxic preconditioning show impair of ovarian function.

BACKGROUND: Cryopreservation of the ovarian tissue has shown promising results. However, there remain controversial issues such as the short half-life of grafts. In this aspect, there are some evidences that preconditioning the ovarian tissue before transplantation is beneficial. OBJECTIVE: To determine the effect of hypoxic preconditioning in vitro on ovarian tissue prior to transplantation. METHODS: Eighteen female adult Wistar rats, were sorted into three experimental groups. Ovaries were maintained in DMEM low glucose serum free at 37°C with 5% CO2, at atmospheric oxigen concentration (normoxia) or 1% O2 (hypoxia) for 16 hours. Oxigen concentration was determined by injection of nitrogen in the incubator. Animals submitted to ovarian transplantation immediately after oophorectomy were the Control Group (C). After this, the ovaries were implanted in the retroperitoneum with nonabsorbable suture and animals evaluated for thirty days after transplantation. Beginning on postoperative (PO) day 11, a daily collection of vaginal smear was carried out. Analyses comprised morphological, morphometric (counting ovarian follicles and corpora lutea) and immunohistochemistry for cleaved caspase-3 (apoptosis). RESULTS: In normoxia and control groups all animals recovered their estrous cycles, while in the hypoxia group, two animals did not ovulate but, among those which did, resumption took longer than in the other groups (p < 0.05). The number of ovarian follicles and corpora lutea decreased significantly in the hypoxia group when compared to the other two groups (p < 0.001) and apoptosis was increased in the few ovarian follicles which remained viable (p < 0.001). CONCLUSION: The hypoxic preconditioning in vitro was not beneficial to the graft and worsened their viability, compromising its functionality or delaying the return of this.

O grau de melhora na função das células progenitoras endoteliais derivadas da medula óssea é dependente do volume de treinamento físico aeróbio / The level of improvement in the function of bone marrow derived endothelial progenitor cells is dependent on the volume of aerobic exercise training

INTRODUÇÃO: A angiogênese muscular esquelética induzida pelo treinamento físico aeróbio (TF) é determinante na melhora da capacidade aeróbia. Entre os fatores envolvidos, as células progenitoras endoteliais (CPE) derivadas da medula óssea são descritas por promoverem o reparo vascular e a angiogênese. Embora o papel do TF sobre os parâmetros das CPE tenha sido investigado, pouco se conhece sobre os efeitos de diferentes volumes de TF sobre a função das CPE da medula óssea, alterações metabólicas e capilarização muscular. OBJETIVO: Testar a hipótese de que o TF melhore a função das CPE da medula óssea, acompanhada por maior capilarização e capacidade oxidativa muscular dependentes do aumento de volume de TF. MÉTODOS: Vinte e uma ratas Wistar foram divididas em três grupos: sedentário controle (SC), treinado protocolo 1 (P1), treinado protocolo 2 (P2). P1: o treinamento de natação consistiu de 60 min, 1x/dia, cinco dias/semana/10 semanas, com 5% de sobrecarga corporal. P2: o mesmo de P1 até a oitava semana, na nona semana os animais treinaram 2x/dia e na 10ª semana 3x/dia. RESULTADOS: O TF promoveu bradicardia de repouso, aumento da tolerância ao esforço, do consumo de oxigênio de pico e da atividade da enzima citrato sintase muscular no grupo P1, sendo estas adaptações mais exacerbadas no grupo P2, indicando que a condição aeróbia foi mais proeminente com este TF. O TF melhorou a função das CPE da medula óssea em P1, sendo ainda maior esta resposta no grupo P2. Em paralelo, observa-se também um aumento no número de capilares dependentes do volume de TF. CONCLUSÃO: Estes resultados sugerem que a medula óssea como o principal reservatório de CPE é influenciada por diferentes volumes de TF, sendo possivelmente responsável pelo maior rendimento físico observado mediante uma maior mobilização endógena de CPE, participantes ativas no processo de angiogênese muscular induzido pelo TF.

INTRODUCTION: Skeletal muscle angiogenesis induced by aerobic exercise training (ET) is crucial in the improvement of the aerobic capacity. The endothelial progenitor cells (EPC) derived from bone marrow have been described for promoting both the vascular repair and angiogenesis. Although the role of the ET on the parameters of the EPC has been investigated, the effect of different volumes of ET on the EPC function in bone marrow, skeletal muscle metabolic alterations and capilarization are unknown. OBJECTIVE: We hypothesized that ET improves the EPC function in bone marrow, accompanied by an increase of skeletal muscle oxidative capacity and angiogenesis dependents of the increase of volume of ET. METHODS: Twenty one Wistar rats were divided into 3 groups: sedentary control (SC), trained protocol 1 (T1) and trained protocol 2 (T2). T1: swimming training consisted of 60 min, 1x/day/10weeks, with 5% body weight load. T2 the same as T1 until 8th week, in the 9th week the rats trained 2x/day and in the 10th week 3x/day. RESULTS: ET promoted resting bradycardia, increase of exercise tolerance, peak oxygen uptake and citrate synthase enzyme activity in the T1 group, being these adaptative responses exacerbate in the P2 group, indicating that the aerobic condition was improved in this group. ET improved the EPC function of the bone marrow in T1, and the response was exacerbed in T2 group. In parallel, an increase in the number of capillaries dependent of ET volume was also observed. CONCLUSION: These findings suggest that the bone marrow as the main reservoir of EPC is influenced by different ET volume, possibly being responsible for the improvement of aerobic performance observed by higher endogenous EPC mobilization, active participants in the process of angiogenesis induced by ET.

Porcine adipose tissue-derived mesenchymal stem cells retain their proliferative characteristics, senescence, karyotype and plasticity after long-term cryopreservation.

We and others have provided evidence that adipose tissue-derived mesenchymal stem cells (ASCs) can mitigate rat cardiac functional deterioration after myocardial ischemia, even though the mechanism of action or the relevance of these findings to human conditions remains elusive. In this regard, the porcine model is a key translational step, because it displays heart anatomic-physiological features that are similar to those found in the human heart. Towards this end, we wanted to establish the cultural characteristics of porcine ASCs (pASCs) with or without long-term cryostorage, considering that allogeneic transplantation may also be a future option. Compared to fresh pASCs, thawed cells displayed 90-95% viability and no changes in morphological characteristics or in the expression of surface markers (being pASCs characterized by positive markers CD29(+); CD90(+); CD44(+); CD140b(+); CD105(+); and negative markers CD31(-); CD34(-); CD45(-) and SLA-DR(-); n = 3). Mean population doubling time was also comparable (64.26±15.11 hours to thawed cells vs. 62.74±18.07 hours to fresh cells) and cumulative population doubling increased constantly until Passage 10 (P10) in the entire cell population, with a small and gradual increase in senescence (P5, 3.25%±0.26 vs. 3.47%±0.32 and P10, 9.6%±0.29 vs. 10.67%±1.25, thawed vs. fresh; SA-ß-Gal staining). Chromosomal aberrations were not observed. In addition, under both conditions pASCs responded to adipogenic and osteogenic chemical cues in vitro. In conclusion, we have demonstrated the growth characteristics, senescence, and the capacity of pASCs to respond to chemical cues in vitro and have provided evidence that these properties are not influenced by cryostorage in 10% DMSO solution.

Adipose tissue-derived stem cells from humans and mice differ in proliferative capacity and genome stability in long-term cultures.

Adipose tissue-derived stem cells (ASCs) are among the more attractive adult stem cell options for potential therapeutic applications. Here, we studied and compared the basic biological characteristics of ASCs isolated from humans (hASCs) and mice (mASCs) and maintained in identical culture conditions, which must be examined prior to considering further potential clinical applications. hASCs and mASCs were compared for immunophenotype, differentiation potential, cell growth characteristics, senescence, nuclear morphology, and DNA content. Although both strains of ASCs displayed a similar immunophenotype, the percentage of CD73(+) cells was markedly lower and CD31(+) was higher in mASC than in hASC cultures. The mean population doubling time was 98.08 ± 6.15 h for hASCs and 52.58 ± 3.74 h for mASCs. The frequency of nuclear aberrations was noticeably lower in hASCs than in mASCs regardless of the passage number. Moreover, as the cells went through several in vitro passages, mASCs showed changes in DNA content and cell cycle kinetics (frequency of hypodiploid, G0/G1, G2/M, and hyperdiploid cells), whereas all of these parameters remained constant in hASCs. Collectively, these results suggest that mASCs display higher proliferative capacity and are more unstable than hASCs in long-term cultures. These results underscore the need to consider specificities among model systems that may influence outcomes when designing potential human applications.

Terapia celular para isquemia cardíaca: efeitos da via de administração, do tempo pós-lesão e do uso biopolímero para a retenção das células e função miocárdica / Cell therapy for ischemic cardiac disease: effect of different routes for cell administration, time post-mi and the use of a fibrin polymer for cardiac cell retention and myocardial function

A terapia celular representa uma abordagem promissora para o tratamento de cardiopatia isquêmica, porém aspectos-chave dessa estratégia permanecem incertos. Neste trabalho avaliamos a eficiência da retenção cardíaca de células da medula óssea marcadas com tecnécio (99m Tc-CMO) transplantadas, de acordo com o tempo após o infarto (1, 2, 3 e 7 dias) e a via de administração dessas células (intravenosa [IV], intraventricular [IC], intracoronariana [ICO] e intramiocárdica [IM]), em ratos submetidos à isquemia-reperfusão cardíaca [I&R]. Após 24 horas, a retenção cardíaca de 99m Tc-CMO foi maior na via IM comparada com a média alcançada pelas demais (6,79% do total injetado vs. 0,53%). O uso de fibrina como veículo para a injeção de células incrementou a retenção em 2.5 vezes (17,12 vs. 6,84%) na via IM. Curiosamente, quando administradas após 7 dias, a retenção de células na via IM alcançou valores próximos dos observados com da matriz de fibrina injetadas 24 h após a I&R (16,55 vs. 17,12%), enquanto que para as demais vias as mudanças foram insignificantes. Nos animais em que as CMO foram administradas por via intramiocárdica 24 horas após a I&R, com ou sem fibrina, observou-se melhora significante do desempenho cardíaco frente ao estresse farmacológico com fenilefrina quando comparados aos controles. Em conjunto, os dados mostram a biodistribuição das células injetadas após a I&R por 4 diferentes vias e 4 intervalos de tempo pós-lesão e indicam que a via IM é a que produz maior retenção cardíaca...

Cell therapy represents a promising approach for ischemic cardiac disease, but key aspects of this strategy remain unclear. We examined the effects of timing and route of administration of bone marrow cells (BMCs) after myocardial ischemia/reperfusion injury (I&R). 99mTc-labeled BMCs were injected by 4 different routes: intravenous (IV), left ventricular cavity (LV), left ventricular cavity with temporal aorta occlusion (LV+) and intramyocardial (IM). The injections were performed 1, 2, 3, or 7 days after infarction. Cardiac retention was higher following the IM route compared to the average values obtained by all other routes (6.79% of the total radioactivity injected vs. 0.53%). Use of a fibrin biopolymer as vehicle during IM injection led to a 2.5-fold increase in cardiac cell accumulation (17.12 vs. 6.84%). Interestingly, the retention of cells administered with culture medium at day 7 post-MI by the IM route was similar to that observed when cells were injected 24 h post-IM using fibrin (16.55 vs 17.12%), whereas no significant changes were observed for the other routes. Cell therapy 24 hs post MI by IM injection, with or without fibrin, resulted in comparable improvement in cardiac function under pharmacological stress compared to control animals. Together, we provide evidence for the biodistribution of 99mTc-labeled BMCs injected post MI by 4 different routes and times post-injury, which shows that the IM rout is the most effective for cardiac cell retention. The use of a fibrin biopolymer further increased cardiac cell retention and its potential long term benefits, beyond 30, on reducing mortality and improving cardiac function deserve to be explored in the future.