Giuseppe Buscemi

Suppressor of Cytokine Signaling 3 Sensitizes Anaplastic Thyroid Cancer to Standard Chemotherapy

We previously showed that cancer cells from papillary, follicular, and anaplastic thyroid carcinomas produce interleukin-4 and interleukin-10, which counteract the cytotoxic activity of conventional chemotherapy through the up-regulation of antiapoptotic molecules. Here, we identify Janus kinase/signal transducers and activators of transcription (STAT) and phosphatidyl inositol 3-kinase (PI3K)/AKT as the down-stream pathways through which these cytokines confer resistance to cell death in thyroid cancer. We found that the absence of suppressors of cytokine signaling (SOCS) molecules allows the propagation of the survival signaling. Exogenous expression of SOCS1, SOCS3, and SOCS5 in the highly aggressive anaplastic thyroid cancer cells reduces or abolishes STAT3 and 6 phosphorylation and PI3K/Akt pathway activation resulting in alteration in the balance of proapoptotic and antiapoptotic molecules and sensitization to chemotherapeutic drugs in vitro. Likewise, exogenous expression of SOCS3 significantly reduces tumor growth and potently enhances the efficacy of chemotherapy in vivo. Our results indicate that SOCS3 regulation of cytokines-prosurvival programs might represent a new strategy to overcome the resistance to chemotherapy-induced cell death of thyroid cancer.

Comparison between local and regional anesthesia in arteriovenous fistula creation.

Purpose Assessment of the effectiveness of Brachial Plexus Block (BPB) via axillary approach compared to regional anesthesia for arteriovenous fistula surgery in patients affected by end-stage renal disease. Methods We compared forty patients randomly divided into two groups. Group A underwent BPB procedure with 15 mL ropivacaine 1% and 10 mL of saline (0.9% NaCl) via axillary approach. Group B received local anesthesia with lidocaine 2%. The forearm blood vessels were assessed by Doppler ultrasonography before and after the intervention. Results BPB performed on Group A was associated with a considerable venous dilation and a significant decrease (48.7%, P<.05) in pulsatility index (PI) measured by Doppler ultrasound. In Group B, PI and venous dilation remained unaltered in the postoperative phase. No complications such as thrombosis or occlusion were encountered among patients who underwent BPB. Conclusions The axillary-approached BPB was more advantageous than local anesthesia. Its effectiveness was because of venous dilation and the decrease in the PI, consequent to the reduction in peripheral resistances and the increase in local blood flow, thus offering an ideal background for fistula creation and short-term patency.

Use of tacrolimus and mycophenolate mofetil as induction and maintenance in simultaneous pancreas-kidney transplantation.

Abstract Clinical trials using quadruple immunosuppression that include the combination of tacrolimus and mycophenolate mofetil have been shown to reduce the incidence of acute rejection episodes in simultaneous pancreas‐kidney (SPK) transplantation. In an attempt to obtain a low rejection rate without antibody induction therapy, we undertook a prospective study of combined tacrolimus and mycophenolate mofetil and steroids as primary immunosuppression for SPK transplantation. In this study, we analyzed 17 patients who received low‐dose intravenous tacrolimus as induction therapy. This was combined with oral tacrolimus, mycophenolate mofetil, and steroids as the primary immunosuppression regimen. There was a significant reduction of empirically and biopsy‐proven rejection with an incidence of 23 % (4 patients). Leukopenia, gastroparesis, and gastrointestinal side‐effects were the cause of discontinuation of mycophenolate mofetil, or low tacrolimus trough level in those patients who developed rejection. All rejection episodes were easy to treat, and none of them required antibody therapy. The combination of tacrolimus with mycophenolate mofetil without antibody induction therapy is effective in preventing early acute rejection. This combination is safe and effective as an alternative immunosuppressive regimen after SPK transplantation.

Abnormal espansion of segmented filamentous bacteria in the gut: a role in pathogenesis of chronic in fiammatory intestinal desease

Human intestinal microbiota create a complex polymicrobial ecology characterized by high population density, wide diversity, and complexity of interactions. Any imbalance of this complex intestinal microbiome, both qualitative and quantitative, might have serious health consequences including an increase in the number and/or alteration in the type of bacteria in the upper gastrointestinal tract, which is referred to as small intestinal bacterial overgrowth (SIBO) syndrome. SIBO is frequently found in persons fulfilling criteria for irritable bowel syndrome (IBS), and the large overlapping of symptoms of these two pathological conditions led some authors to believe that IBS is secondary to SIBO. Interestingly, SIBO is also found in about 25% of patients with Crohns disease. Emerging data show that specific components of gut microbiota, particularly segmented filamentous bacteria (SFB), activate intestinal immunocompetent cells, for example, Th17 cells which have a potential role in pathogenesis of inflammatory intestinal diseases. On the basis of the aforementioned data we postulate that a previously unidentified specific form of SIBO, involving in particular the aberrant expansion of SFB in the gut, could play a role in the onset of chronic intestinal inflammatory diseases through persistent activation of Th17 cells. From this point of view, a successful therapeutic approach to inflammatory intestinal disease patients could be the administration of specific antibiotics directed against SFB to restore the physiological levels of these bacteria in the gut. Furthermore, it could be very useful to identify appropriate laboratory methodologies to monitor the level of SFB in the gut with the aim of preventing their potentially dangerous increase in numbers.

Is secondary hyperparathyroidism-related myelofibrosis a negative prognostic factor for kidney transplant outcome?

Secondary hyperparathyroidism (HP) presenting with hypocalcemia and subsequent increased parathormone (PTH), is mainly identified in patients with chronic renal failure, which has been associated with variable degrees of bone marrow fibrosis. For suitable patients with end-stage renal disease (ESRD), kidney transplantation is recognized as the therapy of choice, being superior to dialysis in terms of quality of life and long-term mortality risk; in this regard interesting data show that increased time on dialysis prior to kidney transplantation is associated with decreased graft and patient survival. In our opinion an important and until now underestimated determinant of graft survival is the proper activity of bone marrow because of the emerging role of hematopoietic stem cells (HSC) in repair of ischemia/reperfusion (IR) damage. We postulate that in ESRD patients, who usually undergo long dialytic treatment, a myelofibrosis caused by an overt secondary HP could drastically decrease the HSC potential for IR damage repair after kidney transplant; this could irremediably lead to a delay in graft function with all related complicances. If the curative role of bone marrow-derived stem cells was confirmed by more data obtained in experimental animal models, it could be possible to try a cellular-based therapeutic approach in the management of ESRD patients which are in waiting list for a kidney transplant.

Isolation and Culture of β-Like Cells From Porcine Wirsung Duct

We sought to develop a protocol to isolate and culture porcine Wirsung duct cells in order to determine their potency to differentiate into insulin-expressing beta-like cells. The porcine Wirsung duct isolated by a surgical microdissection was digested with collagenase P and trypsin to dissociate ductal cells. These elements were cultured in serum-free supplemented media: for 2 weeks. Thereafter the cells were exposed to varying concentrations of glucose (0, 5.6, 17.8, and 25 mmol/L) to induce a beta-like phenotype, as identified by immunohistochemical staining. Cell growth proceeded slowly for the first 2 weeks of culture. After glucose induction for 2 weeks, they formed pancreatic islet-like structures. These cells were stained for the pancreatic ductal cell marker cytokeratin-19 (CK-19) and the pancreatic endocrine markers insulin and glucagon. After the second week, 90% of cells were positive for CK-19. Up to 20.1% of the cells in pancreatic 3-dimensional structures induced by 17.8 mmol/L glucose were positive for insulin, and <3.2%, for glucagon. The positive ratio of immunoreactive staining was dependent on the glucose concentration; 17.8 mmol/L glucose effectively stimulated insulin- and glucagon-secreting cells. We concluded that porcine Wirsung duct cells were capable of proliferation with the potential to differentiate toward beta cells upon glucose induction in vitro.

Use of Intraperitoneal ePTFE Gore Dual-Mesh Plus in a Giant Incisional Hernia After Kidney Transplantation: A Case Report

We evaluated the incidence of and predisposing factors for an incisional hernia after kidney transplantation. Numerous techniques have been used to repair postoperative fascial dehiscences or simple incisional hernias, but no clear treatment exists for giant hernias. Our aim was to obtain (1) a safe procedure to repair a large abdominal defect and reinforce the surrounding, fragile zones and (2) a simple, rapid technique to reduce the operative time. Herein we have described the surgical repair of a giant incisional hernia using intraperitoneal Gore ePTFE dual-mesh plus (Gore-Tex; W. L. Gore, Flagstaff, Ariz, USA) in a 55-year-old man status-post renal transplantation. Total necrosis of distal graft ureter had caused a giant urinoma. The patient was reexplored on day 2 posttransplantation with a primary fascial approximation. Thirty days after transplantation we discovered a large incisional hernia and performed a repair. No drain was used. The patient continued immunosuppressive therapy (cyclosporine, mycophenolate mofetil, prednisolone) and was discharged on postoperative day 4 with no complications. An ultrasonographic follow-up at 1 year revealed the prosthesis to be correctly positioned. Incisional hernia is not rare after renal transplantation but the real incidence is unknown. Immunosuppressive therapy, prolonged pretransplantation dialysis, obesity, and diabetes are probably the major causes of incisional hernias in these patients. Surgical complications of renal transplantation surgery, such as wound hematoma, urinoma, and lymphocele, are the most important predisposing factors for an incisional hernia. The use of intraperitoneal ePTFE dual-mesh is feasible, safe, and easy to repair a large incisional hernia in a kidney transplant patient.

Sistemic calciphylaxis and thrombotic microangiopathy in a kidney transplant patient: Two mixing fatal syndromes?

Abnormalities in calcium and phosphorus metabolism are common and metabolic bone diseases develop often in patients with chronic renal failure (CRF). Effective clinical management includes measures to control phosphorus retention and prevent hyperphosphataemia, to maintain serum calcium concentrations within the normal range and to prevent excess parathyroid hormone (PTH) secretion by the judicious use of vitamin D sterols. Certain of these interventions, however, appear to increase the risk of soft tissue and vascular calcification in patients with End Stage Renal Disease (ESRD), so current therapeutic approaches are thus being re-evaluated in an effort to limit these risks. Patients with calciphylaxis have an extremely high mortality rate, diagnosis requires a high degree of clinical suspicion and the role and extent of parathyroidectomy in the management of this condition remain controversial. In some cases renal transplant patients could suffer from a comorbidity in which vascular function is compromised not only by secondary hyperparathyroidism-related calcification but also by other pathological condition as haemolytic uremic syndrome (HUS), leading to a fatal clinical outcome. We postulate that in these cases a secondary hyperparathyroidism not properly diagnosed in an early phase of the renal disease (probably before the kidney transplant) could cause a vascular calcification which, adding to the pre-existing HUS-related vascular compromission, gave rise to catastrophic clinical consequences. In the management of ESRD patients, in particular in the cases of pre-existing angiopathies, could be therefore crucial the early and proper diagnosis of an alteration of calcium-posphorus metabolism and effort of medicine could be oriented in these cases also towards identification of screening methodologies to undoubtedly assess such a diagnosis.

A complex case of fatal calciphylaxis in a female patient with hyperparathyroidism secondary to end stage renal disease of graft and coexistence of haemolytic uremic syndrome

BACKGROUND Calciphylaxis is a potentially fatal complication of persistent secondary hyperparathyroidism; its cause is still not clear. Unfortunately there is no close relation in severity of clinical picture, serological and pathological alteration. For this reason the prognosis is difficult to establish. Administration of sodium thiosulphate may reduce the precipitation of calcium crystals and improve the general clinical conditions before surgical parathyroidectomy, which seems the only therapeutic approach able to reduce the mortality risk in these patients. METHODS AND RESULTS A 60 year old female patient suffering from End Renal Stage Disease, on haemodialysis from 2001 due to the onset of haemolytic uremic syndrome, underwent a kidney transplant in April 2008. After transplantation there was a recurrence of the haemolytic uremic syndrome, with temporary worsening of the graft. Six months later there was a definite loss of graft and return to dialysis treatment. On April 2010 a severe systemic calciphylaxis related to secondary hyperparathyroidism was diagnosed. The patient underwent parathyroidectomy but, because of the unimproved clinical picture, treatment with sodium thiosulphate was initiated. There was only improvement in cutaneous lesions. The worsening general clinical condition of the patient caused death due to general septic complications. CONCLUSIONS The coexistence of haemolytic uremic syndrome and secondary hyperpathyroidism makes the prognosis poor and, in this case, therapy, which counteracts calcium crystals precipitation, has no effect. Preventive parathyroidectomy can be considered as the only possible treatment.

Dissecting the different biological effects of oncogenic Ras isoforms in cancer cell lines: Could stimulation of oxidative stress be the one more weapon of H-Ras?: Regulation of oxidative stress and Ras biological effects

Ras proteins are small GTPase functioning as molecular switches that, in response to particular extracellular signalling, as growth factors, activate a diverse array of intracellular effector cascades regulating cell proliferation, differentiation and apoptosis. Human tumours frequently express Ras proteins (Ha-, Ki-, N-Ras) activated by point mutations which contribute to malignant phenotype, including invasiveness and angiogenesis. Despite the common signalling pathways leading to similar cellular responses, studies clearly demonstrate unique roles of the Ras family members in normal and pathological conditions and the lack of functional redundancy seems to be explainable, at least in part, by the ability of Ras isoforms to localize in different microdomains to plasma membrane and intracellular organelles. This different intracellular compartmentalization could help Ras isoforms to contact different downstream effectors finally leading to different biological outcomes. Interestingly, it has also been shown that Ha- and Ki-Ras exert an opposite role in regulating intracellular redox status. In this regard we suggest that H-Ras specific induction of ROS (reactive oxygen species) production could be one of the main determinants of the invasive phenotype which characterize cancer cells harbouring H-Ras mutations. In our hypothesis then, while K-Ras (not able to promote oxidative stress) could mainly contribute to cancer progression and invasiveness through activation of MAPK and PI3K, H-Ras-mediated oxidative stress could play a unique role in modulation of intercellular contacts leading to a loss of cell adhesion and eventually also to a metastatic spread.