Cell contact accelerates replicative senescence of human mesenchymal stem cells independent of telomere shortening and p53 activation: roles of Ras and oxidative stress

Mesenchymal stem cells (MSCs) are of great therapeutic potentials due to their multilineage differentiation capabilities. Before transplantation, in vitro culture expansion of MSCs is necessary to get desired cell number. We observed that cell contact accelerated replicative senescence during such process. To confirm the finding as well as to investigate the underlying mechanisms, we cultured both human bone marrow- and umbilical cord blood-derived MSCs under noncontact culture (subculture performed at 60-70% of confluence), or contact culture (cell passage performed at 100% of confluence). It was found that MSCs reached cellular senescence earlier in contact culture, and the doubling time was significantly prolonged. Marked increase of senescence-associated β-galactosidase-positive staining was also observed as a result of cell contact. Cell cycle analysis revealed increased frequency of cell cycle arrest after contact culture. It was noted, however, that the telomere length was not altered during contact-induced acceleration of senescence. Moreover, cell cycle checkpoint regulator P53 expression was not affected by cell contact. Marked increase in intracellular reactive oxygen species (ROS) and a concomitant decrease in the activities of antioxidative enzymes were also observed during contact-induced senescence. Importantly, increased p16(INK4a) following Ras upregulation was found after contact culture. Taken together, cell contact induced accelerated senescence of MSCs, which is telomere shortening and p53 independent. ROS accumulation due to defective ROS clearance function together with Ras and p16(INK4a) upregulation play an important role in contact-induced senescence of MSCs. Overconfluence should therefore be avoided during in vitro culture expansion of MSCs in order to maintain their qualities for clinical application purposes. The contact-induced senescence model reported in this study will serve as a useful model system that allows further study of the molecular mechanisms of senescence in MSCs.     

Endotipsitis as an indication for pediatric liver transplantation

Endotipsitis is a rare but severe complication of transjugular intrahepatic portosystemic shunt (TIPS), a device widely used to treat portal hypertension in adults, but sparsely used in children. We report a case of endotipsitis in a 3‐year‐old child affected of biliary atresia. She underwent a Kasai procedure at 3 months of age but, although the bile flow was restored, she presented upper gastrointestinal bleeding due to portal hypertension 1.5 years later. A TIPS was placed in order to control the hemorrhage. A year after TIPS placement, she started presenting repeated episodes of cholangitis. Blood cultures were positive to Enterobacter cloacae. Even with long antibiotic courses, adjusted to blood cultures, infectious signs were observed after antibiotic withdrawal. Device infection was demonstrated through Positron emission tomography‐Computed tomography scan. The patient was listed for liver transplantation, and intravenous antibiotic treatment was maintained until stent removal during the liver transplant 8 months later. No infectious complications were demonstrated after the surgery. To the best of our knowledge, this is the first case report of endotipsitis described in a pediatric patient.     

Aggressive Epstein‐Barr virus‐negative B‐cell post‐transplant lymphoproliferative disorder in a hepatitis C‐negative liver transplant recipient who received a hepatitis C‐positive graft: Implications for D+/R− hepatitis C virus seroconversion

Post‐transplant lymphoproliferative disorder (PTLD) is an uncommon, but well‐described complication after liver transplantation. Most recently, Hepatitis C virus (HCV) has been implicated in the development of PTLD. A HCV‐negative 62‐year‐old man with autoimmune hepatitis received a HCV nucleic acid amplification test‐positive liver graft from a 73‐year‐old brain‐dead donor (D+/R?). After his recovery from the operation, the patient was treated for HCV and achieved an undetectable viral load. He was readmitted 6 months after transplant with a spontaneous perisplenic hematoma, weight loss, failure to thrive, low‐grade fevers, and abnormal liver function tests. He had a rapid clinical deterioration and expired shortly after admission. His liver biopsy demonstrated EBV‐negative monomorphic B‐cell PTLD. Our case is the first to report an aggressive early‐onset EBV‐negative monomorphic B‐cell PTLD in a HCV D+/R? liver transplant. This case illustrates the paucity of knowledge on HCV seroconversion and its involvement in EBV‐negative monomorphic B‐cell PTLD development.     

The value of repetitive transcranial magnetic stimulation (rTMS) for the treatment of anxiety disorders: an integrative review

Unlike for depression, only few studies are available today investigating the therapeutic effects of repetitive transcranial magnetic stimulation (rTMS) for anxiety disorders. This review aims to provide information on the current research approaches and main findings regarding the therapeutic use of rTMS in the context of various anxiety disorders. Although positive results have frequently been reported in both open and randomized controlled studies, our review of all identified studies indicates that at present no conclusive evidence of the efficacy of rTMS for the treatment for anxiety disorders is provided. Several treatment parameters have been used, making the interpretation of the results difficult. Moreover, sham-controlled research has often been unable to distinguish between response to rTMS and sham treatment. However, there is a limitation in the rTMS methods that likely impacts only the superficial cortical layers. It is not possible to directly stimulate more distant cortical areas, and also subcortical areas, relevant to the pathogenesis of anxiety disorders, though such effects in subcortical areas are thought to be indirect, via trans-synaptic connections. We thus recommend further studies to clearly determine the role of rTMS in the treatment of anxiety disorders. Key advances in combining TMS with neuroimaging technology may aid in such future developments.     

Antiplatelet Agents for Stroke Prevention

Stroke is one of the leading causes of disability and death. Ischemic stroke is a syndrome with heterogeneous mechanisms and multiple etiologies, rather than a singularly defined disease. Approximately one third of ischemic strokes are preceded by another cerebrovascular ischemic event. Stroke survivors are at high risk of vascular events (i.e., cerebrovascular and cardiovascular events), particularly during the first several months after the ischemic event. The use of antiplatelet agents remains the fundamental component of secondary stroke prevention. Based on the available data, antiplatelet agents should be used for patients with noncardioembolic stroke. The use of combination therapy (aspirin plus clopidogrel) has not been proven to be effective or safe to use for prevention of early stroke recurrence or in long-term treatment. There is no convincing evidence that any of the available antiplatelet agents are superior for a given stroke subtype. Currently, the uses of aspirin, clopidogrel, or aspirin combined with extended release dipyridamole are all valid alternatives after an ischemic stroke or transient ischemic attack. However, to maximize the effects of these agents, the treatment should be initiated as early as possible and be continued on a lifelong basis.     

Microscopic cleanliness evaluation of the apical root canal after using calcium hydroxide mixed with chlorhexidine, propylene glycol, or antibiotic paste

This study evaluated cleaning of the dentinal wall after removal of different calcium hydroxide pastes. Sixty-eight single-rooted teeth were prepared using the step-back technique and randomly divided into 4 groups according to medication used: Ca(OH)2 with 0.2% chlorhexidine solution (Group 1), Ca(OH)2 with propylene glycol (Group 2), Ca(OH)2 with antibiotic paste (ciprofloxacin, metronidazole) and distilled water (Group 3), and Ca(OH)2 with antibiotic paste and propylene glycol (Group 4). The samples were stored at 37 °C and 100% relative humidity for 21 days. The medicaments were removed using 5 mL 1% NaOCl, instrumentation with master apical file, 5 mL 1% NaOCl, patency with the K-file #10, ultrasonic instrumentation, and 10 mL 17% EDTA-T. The specimens were analyzed using scanning electron microscopy and chemical analysis. The Kruskal-Wallis (α = 5%) test showed that were no differences between the experimental groups when comparing Ca(OH)2 removal (P = .0951). The chi-square test (α = 5%) indicated a predominance of Ca(OH)2 obstructing dental tubules in all groups. On the basis of the methodology applied, it was concluded that the apical dentine surface remained equally covered by Ca(OH)2, regardless of the vehicle used.     

Carbon dioxide-based nephroscopy during laparoscopic pyeloplasty provides suboptimal view when stones are located in the lower calices

Flexible nephroscopy while using carbon dioxide to insufflate the collecting system has been described as a "good trick" to remove caliceal stones during laparoscopic pyelolithotomy. We found this true only for those calices with undisturbed urine drain into the renal pelvis, while it provided suboptimal view of stones that were located in dependent lower pole calices. We describe how we dealt successfully with a group of lower pole caliceal stones during laparoscopic pyeloplasty.