Molecular cloning and expression of epsilon toxin from Clostridium perfringens type D and tests of animal immunization.

Epsilon toxin produced by Clostridium perfringens types B and D causes enterotoxemia in sheep, goats and calves. Enterotoxemia can cause acute or superacute disease, with sudden death of the affected animal. It provokes huge economic losses when large numbers of livestock are affected. Therapeutic intervention is challenging, because the disease progresses very rapidly. However, it can be prevented by immunization with specific immunogenic vaccines. We cloned the etx gene, encoding epsilon toxin, into vector pET-11a; recombinant epsilon toxin (rec-epsilon) was expressed in inclusion bodies and was used for animal immunization. Serum protection was evaluated and cross-serum neutralization tests were used to characterize the recombinant toxin. To analyze the potency of the toxin (as an antigen), rabbits were immunized with 50, 100 or 200 microg recombinant toxin, using aluminum hydroxide gel as an adjuvant. Titers of 10, 30 and 40 IU/mL were obtained, respectively. These titers were higher than the minimum level required by the European Pharmacopoeia (5 IU/mL) and by the USA Code of Federal Regulation (2 IU/mL). This rec-epsilon is a good candidate for vaccine production against enterotoxemia caused by epsilon toxin of C. perfringens type D.

Phosphorylation and activation of the atypical kinase p53-related protein kinase (PRPK) by Akt/PKB.

p53-related protein kinase (PRPK), the human homologue of yeast Bud32, belonging to a small subfamily of atypical protein kinases, is inactive unless it is previously incubated with cell lysates. Here we show that such an activation of PRPK is mediated by another kinase, Akt/PKB, which phosphorylates PRPK at Ser250. We show that recombinant PRPK is phosphorylated in vitro by Akt and its phospho-form is recognized by a Ser250-phospho-specific antibody; that cell co-transfection with Akt along with wild-type PRPK, but not with its Ser250Ala mutant, results in increased PRPK phosphorylation; and that the phosphorylation of p53 at Ser15, the only known substrate of PRPK, is markedly increased by co-transfection of Akt with wild-type PRPK, but not PRPK dead mutant, and is abrogated by cell treatment with the Akt pathway inhibitor LY294002. Our data disclose an unanticipated mechanism by which PRPK can be activated and provide a functional link between this enigmatic kinase and the Akt signaling pathway.

[Total laparoscopic hysterectomy. Cost analysis]. / Isterectomia totale per via laparoscopica. Analisi dei costi.

BACKGROUND: Hysterectomy performed totally by laparoscopy is still one of the most advanced procedure in endoscopic gynecological surgery. The aim of this study has been to evaluate the cost and the hospital charges of total laparoscopic hysterectomy as compared with those for total abdominal and vaginal hysterectomy. METHODS: In particular, as far as laparoscopic technique is concerned, the cost of linear staplers have been analyzed as compared to disposable and non-disposable instruments for bipolar coagulation. RESULTS: After analysing the data, it has been found that the use of linear staplers involves very high median costs of Lire 2,932,304 ($ 1,650) versus the use of non-disposable instruments of Lire 936,488 ($ 526). The median total hospital charges was of Lire 6,014,448 ($ 3,380) for abdominal hysterectomy, of Lire 4,449,617 ($ 2,500) for vaginal hysterectomy and of Lire 4,078,000 ($ 2,291) for laparoscopic technique with non-disposable supplies. CONCLUSIONS: The results confirm, on the one hand, that bipolar coagulation is a reliable method of hemostasis, and on the other hand that laparoscopic surgery is a technique that can be recommended as an alternative to laparotomy for hysterectomy with unquestionable benefits both for patients and hospital.

[The neuroendocrine and metabolic response to surgical stress]. / La risposta neuro-endocrina e metabolica allo stress chirurgico.

Surgical stress and post-operative pain evoke complex neuroendocrine and tissue responses aiming at defence from injury and recovery of body integrity. Such a reaction appears to be a real "hormonal storm" which is mainly induced by Sympathetic Nervous System and sympatho-adrenomedullary and hypothalamic-pituitary-adrenal systems activation and, in the periphery, by the release of inflammatory mediators at the site of injury. These substances by means of complex regulatory mechanisms and reciprocal interactions induce significant cardiovascular, metabolic, inflammatory and immunologic changes. However, this response may became excessive and contribute to an increase in postoperative morbidity and mortality. Since pain is a major factor in neuroendocrine response triggered by surgical injury every effort must be made to achieve effective intra and post-operative pain control.