Absolute requirement of spermidine for growth and cell cycle progression of fission yeast (Schizosaccharomyces pombe)

Schizosaccharomyces pombe cells that cannot synthesize spermidine or spermine because of a deletion-insertion in the gene coding for S-adenosylmethionine decarboxylase (Deltaspe2) have an absolute requirement for spermidine for growth. Flow cytometry studies show that in the absence of spermidine an overall delay of the cell cycle progression occurs with some accumulation of cells in the G(1) phase; as little as 10(-6) M spermidine is sufficient to maintain normal cell cycle distribution and normal growth. Morphologically some of the spermidine-deprived cells become spherical at an early stage with little evidence of cell division. On further incubation in the spermidine-deprived medium, growth occurs in most of the cells, not by cell division but rather by cell elongation, with an abnormal distribution of the actin cytoskeleton, DNA (4', 6-diamidino-2-phenylindole staining), and calcofluor-staining moieties. More prolonged incubation in the spermidine-deficient medium leads to profound morphological changes including nuclear degeneration.     

Death,treatment decisions and the permanent vegetative state: evidence from families and experts

Some brain injured patients are left in a permanent vegetative state, i.e., they have irreversibly lost their capacity for consciousness but retained some autonomic physiological functions, such as breathing unaided. Having discussed the controversial nature of the permanent vegetative state as a diagnostic category, we turn to the question of the patients’ ontological status. Are the permanently vegetative alive, dead, or in some other state? We present empirical data from interviews with relatives of patients, and with experts, to support the view that the ontological state of permanently vegetative patients is unclear: such patients are neither straightforwardly alive nor simply dead. Having defended this view from counter-arguments we turn to the practical question as to how these patients ought to be treated. Some relatives and experts believe it is right for patients to be shifted from their currently unclear ontological state to that of being straightforwardly dead, but many are concerned or even horrified by the only legally sanctioned method guaranteed to achieve this, namely withdrawal of clinically assisted nutrition and hydration. A way of addressing this distress would be to allow active euthanasia for these patients. This is highly controversial; but we argue that standard objections to allowing active euthanasia for this particular class of permanently vegetative patients are weakened by these patients’ distinctive ontological status.     

Incidence and risk factors for urinary retention following laparoscopic inguinal hernia repair

Background

The incidence of postoperative urinary retention (PUR) has been reported to range from 1% to 22% in patients who have undergone laparoscopic inguinal hernia procedures. The objectives of this study were to determine the incidence of PUR and examine different risk factors that may be associated with the development of PUR in patients who have undergone laparoscopic inguinal hernia procedures.

Methods

A retrospective chart review was performed on 350 patients. Demographics, comorbidities, and operative and postoperative information were collected in patients undergoing laparoscopic inguinal hernia repair by 3 general surgeons from 2007 to 2011. Statistical analysis was done on patient demographics, medical histories, anesthesia notes, and postoperative notes to identify risk factors for the development of urinary retention after laparoscopic inguinal hernia repair.

Results

Three hundred fifty consecutive patients who underwent laparoscopic inguinal hernia repairs were reviewed. Twenty-nine patients developed PUR, an incidence of 8.3%. Age ≥60 years and history of benign prostatic hyperplasia showed significance on multivariate analysis, with odds ratios of 3.0 and 11.0 respectively (P < .05). Anesthesia time ≥2 hours (odds ratio, .75) was a contributing perioperative risk factor but only as an independent risk factor (P < .05).

Conclusions

History of benign prostatic hyperplasia, age ≥60 years, and anesthesia time ≥2 hours were significant independent risk factors for urinary retention after laparoscopic inguinal hernia repair. On multivariate analysis, only history of group and age ≥60 years showed significance. This is 1 of the largest studies to show that the development of PUR in laparoscopic inguinal hernia repair patients is a multifactorial process. Further studies should be conducted to corroborate our findings.     

Life after colectomy for fulminant Clostridium difficile colitis: a 7-year follow up study

Background

The long-term prognosis of patients undergoing colectomy for fulminant Clostridium difficile colitis has not been well studied. The authors present 7-year survival trends in such patients.

Methods

Patients were identified through a pathologic database. Medical records were reviewed and follow-up phone calls made to determine relevant patient history, longevity, and quality of life.

Results

The 61 patients identified had mean and median survival of 18.1 and 3.2 months, respectively, and 1-year, 2-year, 5-year, and 7-year mortality of 68.5%, 79.6%, 88.9%, and 90.7%, respectively. Previous C difficile infection, hypotension, requirement of vasopressors, mental status changes, elevated arterial lactate, decreased platelet counts, intubation, and longer duration on nonoperative therapy were associated with in-hospital mortality. There were no factors correlated with long-term survival.

Conclusions

Patients who require colectomy for fulminant C difficile colitis have a poor prognosis with poor long-term survival and significant morbidity. Although there are several factors associated with in-hospital mortality, there were no factors correlated with long-term survival.     

CAGS AND ACS EVIDENCE BASED REVIEWS IN SURGERY. 49: Is a diverting loop ileostomy and colonic lavage an alternative to colectomy for the treatment of severe Clostridium difficile–associated disease?

Objective: To determine whether a colon-sparing diverting ileostomy with colonic lavage reduces mortality in patients with severe Clostridium difficile–associated disease (CDAD) when compared with colectomy. Design: Retrospective cohort study. Setting & patients: Forty-two patients with diagnosed severe, complicated CDAD who were treated at the University of Pittsburgh Medical Center or VA Pittsburgh Health Care System between June 2009 and January 2011 with diverting loop ileostomy and colonic lavage (warmed polyethylene glycol 3350/electrolyte solution via the ileostomy and postoperative antegrade instillation of vancomycin flushes via the ileostomy). Patients were compared with a historical control group of 42 patients who had a colectomy. Main outcome: Resolution of CDAD. Results: There was no significant difference in age, sex, pharmacologic immuno-suppression and Acute Physiology and Chronic Health Evaluation-II (APACHE-II) scores between the current cohort and historical controls. In the ileostomy group, surgery was performed laparoscopically in 35 patients (83%). This treatment strategy resulted in reduced mortality compared with the historical population (19% v. 50%; odds ratio [OR] 0.24, p = 0.006). Preservation of the colon was achieved in 39 of 42 patients (93%). Conclusion: Loop ileostomy and colonic lavage are an alternative to colectomy in the treatment of severe, complicated CDAD, resulting in reduced morbidity and preservation of the colon.     

The Plasmodium falciparum eIK1 kinase (PfeIK1) is central for melatonin synchronization in the human malaria parasite. Melatotosil blocks melatonin action on parasite cell cycle

Melatonin and its indoles derivatives are central in the synchronization of malaria parasites. In this research, we discovered that melatonin is unable to increase the parasitemia in the human malaria Plasmodium falciparum that lacks the kinase PfeIK1. The PfeIK1 knockout strain is a valuable tool in the screening of indol-related compound that blocks the melatonin effect in wild-type (WT) parasite development. The assays were performed by using flow cytometry with simultaneous labeling for mitochondria viability with MitoTracker Deep Red and nucleus staining with SYBR Green. We found that Melatotosil leads to an increase in parasitemia in P. falciparum and blocks melatonin effect in the WT parasite. Using microscopy imaging system, we found that Melatotosil at 500 nM is able to induce cytosolic calcium rise in transgenic PfGCaMP3 parasites. On the contrary, the compound Triptiofen blocks P. falciparum cell cycle with IC₅₀ 9.76 µM ± 0.6, inhibits melatonin action, and does not lead to a cytosolic calcium rise in PfGCaMP3 parasites. We also found that the synthetic indol-related compounds arrested parasite cycle for PfeIK1 knockout and (WT) P. falciparum (3D7) in 72 hours culture assays with the IC₅₀ values slighting lower for the WT strain. We concluded that the kinase PfeIK1 is central for melatonin downstream signaling pathways involved in parasite cell cycle progression. More importantly, the indol-related compounds block its cycle as an upstream essential mechanism for parasite survival. Our data clearly show that this class of compounds emerge as an alternative for the problem of resistance with the classical antimalarials.     

Development of a whole cell pneumococcal vaccine: BPL inactivation,cGMP production,and stability

Pneumococcal infections impose a large burden of disease on the human population, mainly in developing countries, and the current pneumococcal vaccines offer serotype-specific protection, but do not cover all pathogenic strains, leaving populations vulnerable to disease caused by non-vaccine serotypes. The pneumococcal whole cell vaccine is a low-cost strategy based on non-capsular antigens common to all strains, inducing serotype-independent immunity. Therefore, we developed the process for the cGMP production of this cellular vaccine. Initially, three engineering runs and two cGMP runs were performed in 60-L bioreactors, demonstrating the consistency of the production process, as evaluated by the growth curves, glucose consumption and metabolite formation (lactate and acetate). Cell recovery by tangential filtration was 92 ± 13%. We optimized the conditions for beta-propiolactone (BPL) inactivation of the bacterial suspensions, establishing a maximum cell density of OD₆₀₀ between 27 and 30, with a BPL concentration of 1:4000 (v/v) at 150 rpm and 4 °C for 30 h. BPL was hydrolyzed by heating for 2 h at 37 °C. The criteria and methods for quality control were defined using the engineering runs and the cGMP Lots passed all specifications. cGMP vaccine Lots displayed high potency, inducing between 80 and 90% survival in immunized mice when challenged with virulent pneumococci. Sera from mice immunized with the cGMP Lots recognized several pneumococcal proteins in the extract of encapsulated strains by Western blot. The cGMP whole cell antigen bulk and whole cell vaccine product lots were shown to be stable for up to 12 and 18 months, respectively, based upon survival assays following i.p. challenge. Our results show the consistency and stability of the cGMP whole cell pneumococcal vaccine lots and demonstrate the feasibility of production in a developing country setting.     

Effects of Meal Size on the Release of GLP-1 and PYY After Roux-en-Y Gastric Bypass Surgery in Obese Subjects With or Without Type 2 Diabetes

Changes in gastrointestinal peptide release may play an important role in improving glucose control and reducing body weight following Roux-en-Y gastric bypass (RYGB), but the impact of low caloric intake on gut peptide release post-surgery has not been well characterized. The purpose of this study was to assess the relationships between low caloric intake and gut peptide release and how they were altered by RYGB. Obese females including ten normoglycemic (ON) and ten with type 2 diabetes mellitus (T2DM) (OD) were studied before, 1 week, and 3 months after RYGB. Nine lean, normoglycemic women were studied for comparison. Subjects were given three separate mixed meal challenges (MMCs; 75, 150, and 300 kcal). Plasma glucagon-like peptide 1 (GLP-1) and peptide YY (PYY) were analyzed. Prior to surgery, only minimal increases in GLP-1 and PYY were observed in response to the MMCs. After surgery, the peak GLP-1 concentration was progressively elevated in response to increasing meal sizes. The meal sizes had a statistically significant impact on elevation of GLP-1 incremental areas under the curve (ΔAUC) in both ON and OD at 1 week and 3 months post-surgery visits (p?<?0.05 for all comparisons). The PYY ?AUC was also significantly increased in a meal size-dependent manner in both ON and OD at both post-surgery visits (p?<?0.05 for all comparisons). Meal sizes as small as 75–300 kcal, which cause minimal stimulation in GLP-1 or PYY release in the subjects before RYGB, are sufficient to provide statistically significant, meal size-dependent increases in the peptides post-RYGB both acutely and after meaningful weight loss occurred.     

Chromosome microarray analysis should be offered to all invasive prenatal diagnostic testing following a normal rapid aneuploidy test result

Chromosomal microarray analysis (CMA) has now replaced karyotyping in the analysis of prenatal cases with a fetal structural anomaly, whereas in those pregnancies undergoing invasive prenatal diagnosis with a normal fetal ultrasound, conventional karyotyping is still performed. The aims of this study were to establish the diagnostic yield of CMA in prenatal diagnosis, and to provide new data that might contribute to reconsider current practices. We reviewed 2905 prenatal samples with a normal rapid aneuploidy detection test referred for evaluation by CMA testing. Our study revealed pathogenic and reported susceptibility copy number variants associated with syndromic disorders in 4.8% (n = 138/2905) of cases, being 2.8% (n = 81/2905) the estimated added diagnostic value of CMA over karyotyping. Clinically significant CMA abnormality was detected in 5.4% (107/1975) of the fetuses with ultrasound anomalies and in 1.4% (5/345) of those considered as low-risk pregnancies. Our series shows that in prenatal samples, CMA increases 2-fold the diagnostic yield achieved by conventional karyotyping.