Robotic Site Adjusted Levator Transection for Carcinoma of the Rectum: A Modification of the Existing Cylindrical Abdominoperineal Resection for Eccentrically Located Tumors.

BACKGROUND: Today, extralevator abdominoperineal resection is the standard of care for low rectal cancers with sphincter involvement or location precluding anastomosis. This procedure, while effective from an oncologic point of view, is morbid, with a high incidence of wound complications and genitourinary, and sexual dysfunction. We present a modification of this procedure via a robotic approach, which maintains the radicality while reducing the soft tissue loss and potentially the morbidity. METHODS: Over a 2-year period, five patients (four men and one woman) with eccentric low rectal cancers following neoadjuvant chemoradiation underwent a robot-assisted modified abdominoperineal resection with wide levator transection on the tumor side and conservative levator division on the opposite side. These patients were prospectively followed. Perioperative outcomes, pathologic specimen measures, wound-related problems, and local and systemic recurrences were documented and analyzed. RESULTS: All procedures were successfully completed without conversion. Average body mass index was 32 kg/m2. The mean operative time and blood loss were 370 min and 130 ml, respectively. All specimens had an intact mesorectal envelope with no tumor perforations, and the mean lymph node yield was 16. There were no urinary complications or perineal wound infections. At a median follow-up of 14 months, all patients remain disease-free. CONCLUSIONS: Modified robotic cylindrical abdominoperineal resection with site adjusted levator transection for rectal cancer is an oncologically sound operation in eccentrically located tumors. It maintains the radicality of conventional extralevator abdominoperineal resection, while also reducing the soft tissue loss and thereby potentially the morbidity.

Neurog1 Genetic Inducible Fate Mapping (GIFM) Reveals the Existence of Complex Spatiotemporal Cyto-Architectures in the Developing Cerebellum.

Neurog1 is a pro-neural basic helix-loop-helix (bHLH) transcription factor expressed in progenitor cells located in the ventricular zone and subsequently the presumptive white matter tracts of the developing mouse cerebellum. We used genetic inducible fate mapping (GIFM) with a transgenic Neurog1-CreER allele to characterize the contributions of Neurog1 lineages to cerebellar circuit formation in mice. GIFM reveals Neurog1-expressing progenitors are fate-mapped to become Purkinje cells and all GABAergic interneuron cell types of the cerebellar cortex but not glia. The spatiotemporal sequence of GIFM is unique to each neuronal cell type. GIFM on embryonic days (E) 10.5 to E12.5 labels Purkinje cells with different medial-lateral settling patterns depending on the day of tamoxifen delivery. GIFM on E11.5 to P7 labels interneurons and the timing of tamoxifen administration correlates with the final inside-to-outside resting position of GABAergic interneurons in the cerebellar cortex. Proliferative status and long-term BrdU retention of GIFM lineages reveals Purkinje cells express Neurog1 around the time they become post-mitotic. In contrast, GIFM labels mitotic and post-mitotic interneurons. Neurog1-CreER GIFM reveals a correlation between the timing of Neurog1 expression and the spatial organization of GABAergic neurons in the cerebellar cortex with possible implications for cerebellar circuit assembly.

Cortical contusion injury disrupts olfactory bulb neurogenesis in adult mice.

BACKGROUND: Experimental brain trauma activates quiescent neural stem cells (NSCs) to increase neuronal progenitor cell proliferation in the adult rodent brain. Previous studies have shown focal brain contusion in the form of a unilateral controlled cortical impact (CCI) stimulates NSCs to bilaterally increase neurogenesis in the adult hippocampus. RESULTS: In this study we clarified the bi-lateral effects of a unilateral CCI on proliferation in the subventricular zone (SVZ) NSC niche and on neurogenesis in the olfactory bulb of adult mice. By varying the depth of impact from 1 mm to 2 mm depth, we show CCI to the left somatosensory cortex resulted in graded changes in mouse behavior and cellular pathology in the forebrain. As expected, contusion to the sensorimotor cortex resulted in motor coordination deficits in adult mice. During the first 3 days after injury, CCI increased proliferation in the impacted cortex, deeper striatum and SVZ of the forebrain ipsilateral to the CCI. In each of these regions proliferation was increased with increasing injury severity. At 30 days post-procedure, CCI resulted in a significant reduction in neurogenesis in the olfactory bulb ipsilateral to the CCI. Olfactory avoidance testing indicated disruptions in olfactory bulb neurogenesis were associated with impaired olfactory discrimination in mice post-injury. CONCLUSION: The data demonstrate a focal cortical contusion injury to the left somatosensory cortex disrupts SVZ-olfactory bulb neurogenesis and impairs olfactory discrimination and motor coordination in adult mice.

Review of clinical studies comparing meningococcal serogroup C immune responses induced by MenACWY-TT and monovalent serogroup C vaccines.

Many countries are replacing meningococcal serogroup C (MenC) conjugate vaccines (MCCV) with quadrivalent conjugate (MenACWY) vaccines, such as MenACWY-TT (Nimenrix®). This review examined eight studies comparing MenC immune responses induced by MenACWY-TT and MCCV to determine if these data support these changes. MenC serum bactericidal antibody levels using human (hSBA) or rabbit complement (rSBA) were evaluated at ~1 month postvaccination. Overall, ≥98.4% of infants administered 2 + 1 MenACWY-TT or MCCV schedules had rSBA titers ≥1:8 postprimary and postbooster vaccination; hSBA titers ≥1:8 were similar. In toddlers administered single MenACWY-TT or MCCV doses, ≥97.3% had rSBA titers ≥1:8 postvaccination; percentages with hSBA titers ≥1:8 were higher post-MenACWY-TT. Of children and adolescents receiving primary and booster MenACWY-TT and MCCV, ≥98.6% had rSBA titers ≥1:8; all children receiving MenACWY-TT or MCCV booster had hSBA titers ≥1:8 postdosing. MenC immune responses induced by MenACWY-TT are robust and generally comparable/superior to MCCV, supporting changes to recommendations.

A phase 4 study of the safety of the 13-valent pneumococcal conjugate vaccine in children 6 to 17 years of age in India.

PURPOSE: The 13-valent pneumococcal conjugate vaccine (PCV13) was recently approved in India for the prevention of pneumococcal disease in children aged 6 to 17 years based on global data as well as immunogenicity and safety findings from a phase 3 study. The current phase 4 study in India further evaluated the safety profile of PCV13 in this age group to support the positive benefit-risk profile of PCV13. METHODS: Healthy male and female children aged 6 to 17 years in India were administered a single intramuscular injection of PCV13. Through 7 days after PCV13 administration, local reactions and systemic events were recorded daily by caregivers in an electronic diary. Adverse events (AEs) were collected from the provision of informed consent through 28-42 days postvaccination. RESULTS: One hundred subjects enrolled in and completed the study. After PCV13 vaccination, 73.9% and 57.8% of subjects reported local reactions and systemic events, respectively. The majority of reactogenicity events were mild to moderate in severity, with injection site pain and fatigue the most frequently reported local reaction and systemic event, respectively. Six subjects reported 7 AEs, all of which were considered unrelated to PCV13. One subject reported a serious AE (acute hepatitis), which was considered unrelated to PCV13 and ultimately resolved. No subjects withdrew because of AEs, and there were no deaths. CONCLUSION: PCV13 vaccination was well tolerated with an acceptable safety profile in healthy subjects aged 6 to 17 years in India. This work further supports the safety profile of PCV13 for prevention of pneumococcal disease in this age group in India.

The Decisive Case-Control Study Elaborates the Null Association between ESR1 XbaI and Osteoarthritis in Asians: A Case-Control Study and Meta-Analysis.

(1) Background: The prevalence of knee osteoarthritis (OA) in women is significantly higher than in men. The estrogen receptor α (ERα) has been considered to play a key role due to a large gender difference in its expression. ERα is encoded by the gene estrogen receptor 1 (ESR1), which is widely studied to explore the gender difference in knee OA. Several polymorphisms in ESR1 [PvuII (rs2234693) and BtgI (rs2228480)] were confirmed as the risk factors of OA. However, the evidence of the last widely investigated polymorphism, ESR1 Xbal (rs9340799), is still insufficient for concluding its effect on knee OA. (2) Objective: This study proposed a case-control study to investigate the association between ESR1 Xbal and knee OA. Moreover, a meta-analysis and trial sequential analysis (TSA) were conducted to enlarge the sample size for obtaining a conclusive evidence. (3) Methods: In total, 497 knee OA cases and 473 healthy controls were recruited between March 2015 and July 2018. The Kellgren-Lawrence grading system was used to identify the knee OA cases. To improve the evidence level of our study, we conducted a meta-analysis including the related studies published up until December 2018 from PubMed, Embase, and previous meta-analysis. The results are expressed as odds ratios (ORs) with corresponding 95% confidence intervals (CI) for evaluating the effect of this polymorphism on knee OA risk. TSA was used to estimate the sample sizes required in this issue. (4) Results: We found non-significant association between the G allele and knee OA [Crude-OR: 0.97 (95% CI: 0.78-1.20) and adjusted-OR: 0.90 (95% CI: 0.71-1.15) in allele model] in the present case-control study, and the analysis of other genetic models showed a similar trend. After including six published studies and our case-control studies, the current evidence with 3174 Asians showed the conclusively null association between ESR1 XbaI and knee OA [OR: 0.78 (95% CI: 0.59-1.04)] with a high heterogeneity (I2: 78%). The result of Caucasians also concluded the null association [OR: 1.05 (95% CI: 0.56-1.95), I2: 87%]. (5) Conclusions: The association between ESR1 XbaI and knee OA was not similar with other polymorphisms in ESR1, which is not a causal relationship. This study integrated all current evidence to elaborate this conclusion for suggesting no necessity of future studies.

Safety and immunogenicity of different Clostridioides (Clostridium) difficile vaccine formulations in two early phase randomized studies of healthy adults aged 50-85 years.

BACKGROUND: Two phase 1/phase 2 studies assessed 2 formulations of investigational bivalent Clostridioides (Clostridium) difficile vaccine (QS-21 adjuvanted toxoid and toxoid-alone) in healthy adults 50-85 years of age. METHODS: The QS-21 adjuvanted toxoid vaccine study randomized subjects 3:1 to 100 µg QS-21-containing C difficile vaccine or placebo administered in a shortened-month (Months 0, 1, 3) or day (Days 1, 8, 30) regimen. The toxoid-alone vaccine study randomized subjects 3:3:1 to receive 100 or 200 µg unadjuvanted C difficile vaccine formulation or placebo in Stages 1 and 2 (sentinel cohorts of different age groups), and 3:1 to receive the selected dose of unadjuvanted C difficile vaccine formulation or placebo in Stage 3 (Days 1, 8, 30). Safety was the primary outcome for both studies. Immunogenicity was determined by measuring serum toxin A- and B-specific neutralizing antibodies. RESULTS: In the day regimen, 10 reports across both studies of grade 3 injection site redness postdose 2 triggered predefined stopping rules. Local reactions in both studies were more common among vaccine versus placebo recipients. Injection site pain predominated and was generally mild in severity. Systemic events were infrequent and generally mild-to-moderate in severity. Adverse events were reported by 50.0%-75.0% and 16.7%-50.0% of subjects in the QS-21 and toxoid-alone studies, respectively. Immune responses peaked around Day 37 (shortened-month regimen) or between Day 15 and Month 2 (day regimen) and remained above baseline throughout follow-up. CONCLUSIONS: Both formulations demonstrated robust immunogenicity. Both studies stopped early due to grade 3 injection site redness postdose 2 of the day regimen; neither formulation progressed to later stage development. Instead, an aluminum hydroxide-containing formulation of the vaccine candidate administered at 0, 1, and 6 months, which was safe and immunogenic in phase 1 and 2 studies, advanced to phase 3 studies.

Clinically compatible advances in blood-derived endothelial progenitor cell isolation and reprogramming for translational applications.

The promise of using induced pluripotent stem cells (iPSCs) for cellular therapies has been hampered by the lack of easily isolatable and well characterized source cells whose genomes have undergone minimal changes during their processing. Blood-derived late-outgrowth endothelial progenitor cells (EPCs) are used for disease modeling and have potential therapeutic uses including cell transplantation and the translation of induced pluripotent stem cell (iPSC) derivatives. However, the current isolation of EPCs has been inconsistent and requires at least 40-80 mL of blood, limiting their wider use. In addition, previous EPC reprogramming methods precluded the translation of EPC-derived iPSCs to the clinic. Here a series of clinically-compatible advances in the isolation and reprogramming of EPCs is presented, including a reduction of blood sampling volumes to 10 mL and use of highly efficient RNA-based reprogramming methods together with autologous human serum, resulting in clinically relevant iPSCs carrying minimal copy number variations (CNVs) compared to their parent line.

Alpha-lipoic acid activates eNOS through activation of PI3-kinase/Akt signaling pathway.

BACKGROUND: Lipoic acid (LA) exerts therapeutic effects on cardiovascular diseases. However, the mechanisms underlying these therapeutic effects remain elusive. Endothelial nitric oxide synthase (eNOS) plays a critical role in cardiovascular homeostasis. LA was shown to potently activate PI3-kinase/Akt pathway, and the latter is critical in the regulation of eNOS activity. In the present study, we test the hypothesis that LA improves endothelial function through PI3-kinase/Akt-mediated eNOS activation. METHODS AND RESULTS: Western blot analysis showed that LA time- and dose-dependently induced phosphorylation of Akt and eNOS in human umbilical vein endothelial cells (HUVECs). Both PI3-kinase and Akt inhibitors abolished LA-induced eNOS phosphorylation, indicating that LA induces eNOS phosphorylation through the PI3-kinase/Akt pathway. This increase in eNOS phosphorylation was paralleled by an increase in NO release by HUVECs, supporting its relevance in eNOS activity regulation. Myograph analysis revealed that LA relaxed phenylephrine-induced contraction. Endothelium removal and NOS inhibition by L-NAME abolished this vasodilator action of LA, and Akt but not AMPK inhibition significantly reduced the vasodilator action of LA, indicating that it is mediated by PI3-kinase/Akt pathway-dependent activation of eNOS. Consistent with in vitro results, intraperitoneal injection with LA significantly increased plasma nitrite and nitrate levels in C57Bl/6j mice. CONCLUSIONS: LA activates eNOS through a PI3-kinase/Akt signaling pathway-dependent mechanism, offering a potential molecular basis for the therapeutic effects of LA on cardiovascular diseases.

Efficient Reprogramming of Human Fibroblasts and Blood-Derived Endothelial Progenitor Cells Using Nonmodified RNA for Reprogramming and Immune Evasion.

mRNA reprogramming results in the generation of genetically stable induced pluripotent stem (iPS) cells while avoiding the risks of genomic integration. Previously published mRNA reprogramming protocols have proven to be inconsistent and time-consuming and mainly restricted to fibroblasts, thereby demonstrating the need for a simple but reproducible protocol applicable to various cell types. So far there have been no published reports using mRNA to reprogram any cell type derived from human blood. Nonmodified synthetic mRNAs are immunogenic and activate cellular defense mechanisms, which can lead to cell death and inhibit mRNA translation upon repetitive transfection. Hence, to overcome RNA-related toxicity we combined nonmodified reprogramming mRNAs (OCT4, SOX2, KLF4, cMYC, NANOG, and LIN28 [OSKMNL]) with immune evasion mRNAs (E3, K3, and B18R [EKB]) from vaccinia virus. Additionally, we included mature, double-stranded microRNAs (miRNAs) from the 302/367 cluster, which are known to enhance the reprogramming process, to develop a robust reprogramming protocol for the generation of stable iPS cell lines from both human fibroblasts and human blood-outgrowth endothelial progenitor cells (EPCs). Our novel combination of RNAs enables the cell to tolerate repetitive transfections for the generation of stable iPS cell colonies from human fibroblasts within 11 days while requiring only four transfections. Moreover, our method resulted in the first known mRNA-vectored reprogramming of human blood-derived EPCs within 10 days while requiring only eight daily transfections.