Chemical characterization (LC-MS-ESI), cytotoxic activity and intracellular localization of PAMAM G4 in leukemia cells.

Generation 4 of polyamidoamine dendrimer (G4-PAMAM) has several biological effects due to its tridimensional globular structure, repetitive branched amides, tertiary amines, and amino-terminal subunit groups liked to a common core. G4-PAMAM is cytotoxic due to its positive charges. However, its cytotoxicity could increase in cancer cells due to the excessive intracellular negative charges in these cells. Furthermore, this work reports G4-PAMAM chemical structural characterization using UHPLC-QTOF-MS/MS (LC-MS) by electrospray ionization to measure its population according to its positive charges. Additionally, the antiproliferative effects and intracellular localization were explored in the HMC-1 and K-562 cell lines by confocal microscopy. The LC-MS results show that G4-PAMAM generated multivalent mass spectrum values, and its protonated terminal amino groups produced numerous positive charges, which allowed us to determine its exact mass despite having a high molecular weight. Additionally, G4-PAMAM showed antiproliferative activity in the HMC-1 tumor cell line after 24 h (IC50 = 16.97 µM), 48 h (IC50 = 7.02 µM) and 72 h (IC50 = 5.98 µM) and in the K-562 cell line after 24 h (IC50 = 15.14 µM), 48 h (IC50 = 14.18 µM) and 72 h (IC50 = 9.91 µM). Finally, our results showed that the G4-PAMAM dendrimers were located in the cytoplasm and nucleus in both tumor cell lines studied.

Effects of increased apical enlargement on the amount of unprepared areas and coronal dentine removal: a micro-computed tomography study.

AIM: To evaluate the effects of progressive apical enlargement on the amount of unprepared root canal surface area and remaining dentine thickness. METHODOLOGY: The root canals of 30 extracted mandibular incisors with Vertucci's type I configuration were instrumented with rotary HyFlex CM instruments (Coltene-Whaledent, Altstätten, Switzerland) up to 4 instruments larger than the first one that bound at the working length (WL). Teeth were scanned in a micro-computed tomography (micro-CT) device before canal preparation and after instrumentation with the 2nd, 3rd and 4th larger instruments. The amount of unprepared surface area in the full canal or in the apical 4 mm as well as the remaining dentine thickness at 10 mm from the WL were calculated and compared. The general linear model for repeated measures adjusted by Bonferroni's post hoc test was used for statistic analysis. RESULTS: There was a significant reduction in the amount of unprepared areas after each increase in preparation size (P < 0.01). This was observed for both the full canal length and the 4-mm apical segment. The amount of remaining dentine was also significantly reduced after each file size (P < 0.01). However, dentine thickness always remained greater than 1 mm, even after using the largest instrument. CONCLUSION: Apical preparations up to 4 instruments larger than the first one to bind at the WL caused a significant progressive reduction in the unprepared canal area.

What happens to unprepared root canal walls: a correlative analysis using micro-computed tomography and histology/scanning electron microscopy.

AIM: To microscopically examine the cleanliness of root canal walls that remained unprepared as revealed by micro-CT. METHODOLOGY: The root canals of 10 freshly extracted mandibular premolars with necrotic pulps and apical periodontitis along with the mesiobuccal canals of 11 mandibular molars with vital pulps were prepared using Reciproc instruments R40 and R25, respectively, and 2.5% sodium hypochlorite irrigation. Specimens were scanned in micro-CT before and after preparation, and the unprepared areas were identified. The outer root surface corresponding to the untouched areas was marked on each root third to guide further analysis using histological (for teeth with vital pulps) and scanning electron microscopic (SEM; for necrotic teeth) examination. In the teeth with vital pulps, the root canal area occupied by tissue remnants was calculated. In SEM analysis of teeth with necrotic pulps, scores were attributed for the amount of debris on the untouched areas. RESULTS: The proportion of unprepared areas in the mesiobuccal molar canals was 18.1% and 9.6% over the full canal length and apical canal, respectively. In premolars, corresponding figures were 34.6% and 17.6%, respectively. Histological analysis of canals with vital pulps revealed tissue remnants over the untouched walls almost exclusively in the apical canal. SEM analysis of the canals with necrotic pulps revealed debris along the untouched walls in all root canal thirds. CONCLUSION: The areas that remain untouched by Reciproc instruments used with 2.5% NaOCl irrigation as revealed by micro-CT analysis were usually covered with debris, in the form of pulp tissue remnants, bacteria and dentine chips, especially in the apical root canal.

Recent advances in stem and germ cell research: implications for the derivation of pig pluripotent cells.

Pluripotent stem cells have the unique capacity to contribute to all the tissues of an adult animal after transfer into a host embryo. How pluripotency is acquired during early development and how it is maintained in stem cells have attracted the interest of many scientists for over three decades. Much progress in our understanding of how stem cells arise in culture and the signals required for homoeostasis has enabled the derivation of pluripotent cells in multiple species. Here, we discuss recent developments in stem cell biology that will impact the generation of pluripotent cells from different embryonic origins and will contribute to increase our capacity for generating transgenic animals.

Immunoneuroendocrine alterations in patients with progressive forms of chronic Chagas disease.

We studied the features of parallel immunoneuroendocrine responses in patients with different degrees of chronic Chagas myocarditis (indeterminate, mild/moderate or severe). A systemic inflammatory scenario was evident in patients with severe myocarditis compared to healthy subjects. This was paralleled by a disrupted activation of the hypothalamus-pituitary-adrenal axis, characterized by decreased concentrations of dehydroepiandrosterone-sulfate (DHEA-s) and an unbalanced cortisol/DHEA-s ratio, reinforcing the view that severe Chagas disease is devoid of an adequate anti-inflammatory milieu, likely involved in pathology. Our study constitutes the first demonstration of neuroendocrine disturbances, in parallel to a systemic inflammatory profile, during progressive human Chagas disease.

Renal transplantation program in Cuba.

Kidney transplantation has been performed in Cuba since 1970. In 1979, compatible living-related donors were introduced into our renal transplantation program. There are 43 hospitals distributed around the country with a multidisciplinary group that attends cadaveric donors with encephalic death. The donor rate in Cuba oscillates between 15 and 18 per million; 90% of them are from cadaveric donors. This program includes 47 dialysis centers throughout the country with 2300 patients as supported by a National Coordinating Center at The Nephrology Institute. Cuba is one of the first countries in our region with this experience.

Short treatment with the tumour necrosis factor-alpha blocker infliximab diminishes chronic chagasic myocarditis in rats without evidence of Trypanosoma cruzi reactivation.

Tumour necrosis factor (TNF)-alpha is crucial for resistance to Trypanosoma cruzi acute infection, but there is scant information on its role during the chronic phase. To address this issue, we analysed whether a short treatment with a TNF-alpha blocker affected the course and characteristics of chronic disease in a rat experimental model of T. cruzi infection. An anti-TNF-alpha agent (infliximab) was administered during the chronic phase for a period of 4 weeks (3 mg/kg/week), while control infected rats were inoculated with saline physiological solution. Search for parasites yielded non-successful results in all infected groups, irrespective of treatment. Nevertheless, the presence of T. cruzi kDNA in heart tissue was detected in infected and infected plus treated animals. Because infliximab might induce changes in the anti-parasite cytokine response, circulating levels of interleukin (IL)-10, interferon-gamma and nitric oxide were evaluated. An increase in IL-10 levels was observed only in the infected group treated with the anti-TNF-alpha blocker compared to the remaining groups (P < 0.05). A clear attenuation of histological damage associated with a diminution of cardiac TNF-alpha mRNA expression was observed in the infected and treated animals compared to the infected and non-treated group. Blocking of TNF-alpha during a relatively short period in chronically infected rats did not lead to evident parasite reactivation but reduced myocarditis severity significantly, indicating a role of this cytokine in the pathogenesis of chronic myocardial damage.

Improved outcome of Trypanosoma cruzi infection in rats following treatment in early life with suspensions of heat-killed environmental Actinomycetales.

The well-established model of Chagas' disease in "l" rats was used to evaluate the effects of three injections of heat-killed Gordonia bronchialis, Rhodococcus coprophilus or saline on Trypanosoma cruzi parasitaemia and acute and chronic myocarditis, sequelae of the infection. Two vaccinating injections were given prior to challenge with T. cruzi, and the third, immunotherapeutic, injection was given 7 days after challenge. Treatment with either actinomycete significantly reduced acute parasitaemia (p<0.04), modified cellular infiltration during acute myocarditis and limited chronic myocarditis (p<0.03) in comparison with the saline-treated control animals. Immunological investigations showed that both bacterial preparations achieved their results through different mechanisms. The relevance of our findings to human Chagas' disease is discussed.

Hemodynamic and oxidative stress profile after exercise in type 2 diabetes.

This study investigated the effect of an acute bout of exercise (>85% VO2Max) on biochemical, hemodynamic and oxidative stress variables in sedentary and physically active subjects with type 2 diabetes (T2D). Blood measurements were taken before and after a treadmill test on 12 sedentary non-diabetes subjects (ND), 12 sedentary type 2 diabetes (T2S) and 9 physically active T2D subjects (T2DA). T2DS subjects before and after the treadmill test showed a higher plasma glucose (123.2 +/- 19.0 mg/dL versus 108.9 +/- 16.8 mg/dL, p < 0.001), HbA1C (8.7 +/- 2.4% versus 7.3 +/- 1.2%, p < 0.001) and body fat% (21.3 +/- 5.7% versus 34.6 +/- 4.5%, p < 0.001) than T2DA subjects. T2DA had higher VO2Max (37.7 +/- 3.5 versus 29.5 +/- 3.2, p < 0.05), time on treadmill (22.3 +/- 2.1 min versus 16.1 +/- 2.1 min, p < 0.05), hemoglobin (17.9 +/- 0.9 g/dL, p < 0.05) and lower blood pressure levels in comparison to ND and T2DS subjects. Thiobarbituric acid substances (TBARS) in T2DS were higher than in T2DA subjects (0.27 +/- 0.1 nmol/mL versus 0.21 +/- 0.1 nmol/mL, p < 0.05). Glutathione (GSH) levels were similar among the groups. Physically active type 2 diabetes subjects had a more favorable biochemical, hemodynamic and oxidative stress profile than sedentary subjects. The coexistence of a poor cardiopulmonary performance and high oxidative stress environment can determine a profile of high risk for serious cardiovascular events in patients with diabetes.

Deficient control of Trypanosoma cruzi infection in C57BL/6 mice is related to a delayed specific IgG response and increased macrophage production of pro-inflammatory cytokines.

Earlier work in Trypanosoma cruzi-infected C57BL/6 and BALB/c mice revealed an acute disease, of lethal outcome in the former group and lesser severity in BALB/c mice. Fatal course was not accompanied by an increased parasite load, but by a substantial imbalance between pro- and anti-inflammatory cytokine serum levels. To better characterise the mechanisms allowing the host to restrain the infection, we have now studied the specific IgG production and in vitro behaviour of peritoneal macrophages (PMs) when exposed to T. cruzi. BALC/c mice displayed higher serum levels of specific immunoglobulins in the first weeks of acute infection. In vitro infected PMs showed no between-group differences in the number of intracellular parasites, although TNFalpha levels were significantly higher in culture supernatants from C57BL/6 mice. Because an LPS-based pretreatment (desensitisation protocol followed by a sublethal LPS dose) reduced disease severity of C57BL/6 mice, we next explored the features of the in vitro infection in PMs from mice subjected to such protocol. PMs from LPS-pretreated mice had a decreased production of TNFalpha and IL-1beta, becoming more permissive to parasite replication. It is concluded that deficient control of T. cruzi infection in C57BL/6 mice may also involve a less satisfactory specific IgG response and increased TNFalpha production by PMs. Improved disease outcome in LPS-pretreated mice may be associated with the reduced inflammatory cytokine production by PMs, but the impaired ability of these cells to control parasite growth suggests that compensatory mechanisms are operating in the in vivo situation.

Obesity adversely affects the outcome of antireflux operations.

BACKGROUND: We hypothesized that obesity was associated with long-term failure of antireflux procedures, and that in obese patients antireflux operations were easier to perform via thoracotomy, and therefore likely to have a higher success rate than transabdominal (laparoscopic or open) antireflux procedures. The aims of this study was to determine the impact of obesity on the success of antireflux operations, and to compare the success rates of transthoracic and laparoscopic approaches in obese patients with gastroesophageal reflux. METHODS: The records of 224 consecutive patients undergoing antireflux surgery by two surgeons in a university-based tertiary care center were reviewed and patients contacted for follow-up assessment. The patients were classified into groups based on the type of operation performed and the calculated body mass index (BMI): normal (BMI 30). Recurrences were documented by symptoms responsive to acid-suppressive medication and radiologic or pH probe studies. RESULTS: Among the 224 patients included in this study, 187 underwent laparoscopic Nissen fundoplications (LNF) and 37 underwent Belsey Mark IV(BM4) procedures. The mean follow-up period was 37 months. The three groups included 89 (39.7%) patients classified as having normal weight, 87 (38.8%) as overweight and 48 (21.4%) as obese. Normal, overweight, and obese patients were similar in terms of age, gender, hiatal hernia size, degree of esophagitis, and comorbid conditions. A total of 26 recurrences occurred, giving an overall recurrence rate of 11.6%. There were 4 recurrences in the normal group (4.5%), 7 in the overweight group (8.0%; p not significant vs normal), and 15 in the obese group (31%; p < 0.001 vs normal; p <.001 vs overweight). The recurrence rate was similar between LNF and BM4 in each BMI subgroup, although in aggregate, the recurrence rate after BMW was greater than after LNF (10/37 vs 16/187; p < 0.02). CONCLUSIONS: Obesity adversely affects the long-term success of antireflux operations. Although athoracotomy provides optimal exposure of the hiatal structures in obese patients, a transthoracic approach was associated with a higher recurrence rate than LNF. Given the high failure rate of antireflux operations in obese patients, intensive efforts at sustained weight loss should be made before consideration of surgery.

[Helicobacter pylori infection in a group of HIV/AIDS patients]. / Infección por Helicobacter pylori en un grupo de pacientes VIH/SIDA.

The prevalence of H. pylori infection in AIDS patients has been controversial. A. controlled prospective and descriptive study was conducted in a group of 64 HIV-infested patients. H. pylori detection was carried out by endoscopic study during which two biopsy samples of gastric antrum and body were taken for the histologic study and other 2 samples of gastric antrum and body for urease testing. H.pylori infection prevalence was 56.3%. The comparison group was composed by 110 serologically HIV-negative patients, with an infection prevalence rate of 77.2%. In the HIV-AIDS group, the frequency of gastric ulcer was 1,5% and no duodenal ulcer was found whereas the comparison group showed a frequency of gastric and duodenal ulcers of 1.8 and 15.5% respectively. HIV+/AIDS patients in category I exhibited an H. pylori infection prevalence of 68.7%, category II, 39.2% and category III, 21,0%. A reversed association was observed between the level of immune deterioration and the prevalence of H. pylori infection.

[Stomas in pregnancy, clinical case and review of the literature]. / Estomas durante el embarazo, caso clínico y revisión de la literatura.

Stomas and pregnancy is an uncommon event and the literature in this regard is scarce, this poses significant concern on its management. Among the etiology we found the ulcerative colitis, trauma, and rectovaginal fistula, etc. The management should include a perinatologist and a specialist in colon and rectum. We should be familiarized with the potential complication as the intestinal obstruction, stoma prolapse, narrowing of the stoma and bleeding. The route delivery should be vaginal and the c-section is reserved for obstetric indications. The patient must receive education regarding stoma complications, and how to copy to live with a stoma.

[Neumopericardium and spontaneous post partum subcutaneus emphysema. A case report and literature review]. / Neumopericardio y enfisema subcutáneo espontáneos posparto. Presentación de un caso y revision de la literatura.

The barotrauma cases related to childbirth are an uncommon event, especially the neumopericardium. This entity must be suspected in a patient with subcutaneuos emphysema and confirmed by chest X-ray. The management is based on closed observation, support measures and antibiotic to prevent infection. The radiological follow up is important to evaluate the resolution of this problem. The patient must be followed with caution if the neumopericardium increases because this could lead to a cardiac tamponade with fatal consequences. The description of a case diagnosed and managed in our hospital is done herein which evolution was satisfactory towards resolution of the pneumopericardium without the need of further intervention.

[Prescription of exercise in the diabetic patient]. / Acerca de la prescripción de ejercicio en el paciente diabético.

There is a broad epidemiologic evidence related with the increase of the incidence and prevalence of diabetes mellitus type 2 throughout the world, especially in Western populations, at rates considered epidemic. Cerebrovascular disease and myocardial infarction are two of the most important complications of this disease and they have very high social and economic consequences. Treatment of this disease is directed good metabolic control, diminishing toward obtain co-morbidity, and reducing acute and chronic diabetic complications. Diet, hypoglycemic drugs or insulin, and programs of physical activity are used for this purpose. This document presents some thoughts on the difficulties related with the prescription of physical exercise in the diabetic patient and a brief proposal to solve those problems in the clinical practice.

[Rectal catheter decompression in colonic pseudoobstruction (Ogilvie syndrome). Report of a case and review of the literature]. / Descompresión con sonda rectal en la pseudoobstrucción colónica (síndrome de Ogilvie). Informe de un caso y revisión de la literatura.

The Ogilvie's Syndrome is a major surgical complication. It is defined as a severe dilatation of the colon, the cecum, ascendant and transverse, without the presence air at the distal colon following abdominal surgery. The causes are unknown. We described a clinical case of patient delivered by C-S who developed this syndrome and a review of the literature. We emphasized on the importance of the early diagnosis by the use of the traditional abdomen X-Rays and we described a relatively new approach for this patients through the decompression by colonoscopy using a Levin's catheter successfully. This is an alternative minimum-invasive treatment which could be of a great benefit for the patients with this disease.

SpoIIB localizes to active sites of septal biogenesis and spatially regulates septal thinning during engulfment in bacillus subtilis.

A key step in the Bacillus subtilis spore formation pathway is the engulfment of the forespore by the mother cell, a phagocytosis-like process normally accompanied by the loss of peptidoglycan within the sporulation septum. We have reinvestigated the role of SpoIIB in engulfment by using the fluorescent membrane stain FM 4-64 and deconvolution microscopy. We have found that spoIIB mutant sporangia display a transient engulfment defect in which the forespore pushes through the septum and bulges into the mother cell, similar to the situation in spoIID, spoIIM, and spoIIP mutants. However, unlike the sporangia of those three mutants, spoIIB mutant sporangia are able to complete engulfment; indeed, by time-lapse microscopy, sporangia with prominent bulges were found to complete engulfment. Electron micrographs showed that in spoIIB mutant sporangia the dissolution of septal peptidoglycan is delayed and spatially unregulated and that the engulfing membranes migrate around the remaining septal peptidoglycan. These results demonstrate that mother cell membranes will move around septal peptidoglycan that has not been completely degraded and suggest that SpoIIB facilitates the rapid and spatially regulated dissolution of septal peptidoglycan. In keeping with this proposal, a SpoIIB-myc fusion protein localized to the sporulation septum during its biogenesis, discriminating between the site of active septal biogenesis and the unused potential division site within the same cell.