Hypoxia augments MHC class I antigen presentation via facilitation of ERO1‐α‐mediated oxidative folding in murine tumor cells

To establish an effective cancer immunotherapy, it is crucial that cancer cells present a cancer‐specific antigen in a hypoxic area, a hallmark of the tumor microenvironment. Here, we show the impact of hypoxia on MHC class I antigen presentation in vitro and in vivo in murine tumors. Activation of antigen‐specific CTLs by tumor cells that had been pre‐incubated under a condition of hypoxia was enhanced compared with that by tumor cells pre‐incubated under a condition of normoxia. Cell surface expression of MHC class I‐peptide complex on the tumor cells was increased under a condition of hypoxia, thereby leading to higher susceptibility to specific CTLs. We show that the hypoxia‐inducible ER‐resident oxidase ERO1‐α plays an important role in the hypoxia‐induced augmentation of MHC class I‐peptide complex expression. ERO1‐α facilitated oxidative folding of MHC class I heavy chains, thereby resulting in the augmentation of cell surface expression of MHC class I‐peptide complex under hypoxic conditions. These results suggest that since the expression of MHC class I‐peptide complex is augmented in a hypoxic tumor microenvironment, strategies for inhibiting the function of regulatory T cells and myeloid‐derived suppressor cells and/or immunotherapy with immune checkpoint inhibitors are promising for improving cancer immunotherapy.     

Media conditioned by retinal pigment epithelial cells suppress the canonical Wnt pathway

Retinal pigment epithelial (RPE) cells play critical roles in the maintenance of visual function, partly by secreting various biologically active factors that modulate the intraocular environment. Recent studies suggest involvement of Wnt proteins secreted by RPE cells in the pathogenesis of photoreceptor degeneration. In the present study, we examined, via the luciferase assay, the effect of media conditioned by RPE cells (RPE-CM) on activity of the canonical Wnt pathway in vitro. We isolated primary RPE cells from Long-Evans rats at P6-P9. In culture, these cells formed a monolayer with polygonal cell morphology and demonstrated repigmentation at confluency and immunoreactivity for ZO-1, a marker for tight junctions. To evaluate the effect of RPE-CM on the canonical Wnt pathway, we replaced the culture media of COS-7 cells transfected with (Tcf)(7)LUC, a multimeric Tcf-responsive element luciferase reporter construct, with RPE-CM and measured luciferase activity with or without Wnt3a or SB216763, a specific GSK3 inhibitor. RPE-CM did not enhance basal or Wnt3a-induced (Tcf)(7)LUC activity; instead, this activity decreased by 60%. RPE-CM also reduced SB216763-induced (Tcf)(7)LUC activity by 65%, which suggests that the inhibitory effect of RPE-CM is probably due to intracellular crosstalk rather than extracellular antagonism. RPE cells may thus be able to modulate the intraocular environment by regulating the canonical Wnt pathway.     

The Role of Patient–Provider Communication in Engagement and Re-engagement in HIV Treatment in Bamako,Mali: A Qualitative Study

Mounting evidence in sub-Saharan Africa suggests poor patient-provider communication (PPC) negatively impacts patient engagement (retention in care and adherence to medication) in antiretroviral therapy (ART) programs. In Bamako, Mali, where 36% of ART patients are lost to follow-up within 12 months of initiating treatment, we aimed to define features of positive PPC according to patient values and explore the mechanisms by which these features may sustain engagement and re-engagement according to patient and provider experiences. We conducted 33 in-depth interviews and 7 focus groups with 69 patients and 17 providers in five ART clinics. Regarding sustaining engagement, participants highlighted “establishing rapport” as a foundational feature of effective PPC, but also described how “responding to emotional needs”, “eliciting patient conflicts and perspective” and “partnering to mitigate conflicts” functioned to address barriers to engagement and increase connectedness to care. Patients who had disengaged felt that “communicating reacceptance” may have prompted them re-engage sooner and that tailored “partnering to mitigate conflicts” would be more effective in sustaining re-engagement than the standard adherence education providers typically offer. Optimizing provider skills related to these key PPC features may help maximize ART patient engagement, ultimately improving health outcomes and decreasing HIV transmission in sub-Saharan Africa.     

Orbital seeding of mesenchymal stromal cells increases osteogenic differentiation and bone-like tissue formation

In bone tissue engineering (TE), an efficient seeding and homogenous distribution of cells is needed to avoid cell loss and damage as well as to facilitate tissue development. Dynamic seeding methods seem to be superior to the static ones because they tend to result in a more homogeneous cell distribution by using kinetic forces. However, most dynamic seeding techniques are elaborate or require special equipment and its influence on the final bone tissue-engineered construct is not clear. In this study, we applied a simple, dynamic seeding method using an orbital shaker to seed human bone marrow–derived mesenchymal stromal cells (hBMSCs) on silk fibroin scaffolds. Significantly higher cell numbers with a more homogenous cell distribution, increased osteogenic differentiation, and mineral deposition were observed using the dynamic approach both for 4 and 6 hours as compared to the static seeding method. The positive influence of dynamic seeding could be attributed to both cell density and distribution but also nutrient supply during seeding and shear stresses (0.0-3.0 mPa) as determined by computational simulations. The influence of relevant mechanical stimuli during seeding should be investigated in the future, especially regarding the importance of mechanical cues for bone TE applications. Our results highlight the importance of adequate choice of seeding method and its impact on developing tissue-engineered constructs. The application of this simple seeding technique is not only recommended for bone TE but can also be used for seeding similar porous scaffolds with hBMSCs in other TE fields.     

In situ label‐free cell viability assessment of nucleus pulposus tissue

Regenerative medicine approaches aiming at treating degenerating intervertebral discs, a major cause of back pain, are increasingly tested in ex‐vivo disc explant models mimicking in‐vivo conditions. For assessing the efficacy of regenerative therapies, cell viability is commonly measured requiring specific labels to stain cells. Here, we demonstrate and evaluate how cellular auto‐fluorescence can be utilized to non‐invasively assess viability in disc tissue in‐situ using label‐free two‐photon microscopy. Live and dead bovine disc cells (0% and 100% cell viability) from the nucleus pulposus were seeded into collagen gels and auto‐fluorescence was characterized. Subsequently, nucleus pulposus explants were cultured for 6 days in media with different glucose supplementation (0, 0.25, 0.5, and 1 g/L) to induce different degrees of cell death. Then, samples were split and viability was assessed using label‐free two‐photon microscopy and conventional staining. Results show that live and dead nucleus pulposus cells systematically emit auto‐fluorescent light with distinct characteristics. Cell viability values obtained with label‐free microscopy did not significantly differ from those acquired with staining. In summary, monitoring auto‐fluorescence facilitates accurate cell viability assessment in nucleus tissue requiring no additional dyes. Thus, this technique may be suitable for pre‐clinical testing of regenerative therapies in nucleus pulposus cultures. © 2014 Orthopaedic Research Society. Published by Wiley Periodicals, Inc. J Orthop Res 32:545–550, 2014.     

Risk factors for incisional surgical site infections in elective surgery for colorectal cancer: focus on intraoperative meticulous wound management

Purpose

An incisional surgical site infection (I-SSI) is a frequently observed complication following colorectal surgery. Intraoperative wound management is one of the most important factors that determine the incidence of postoperative I-SSI. The purpose of this study was to assess the impact of the methods used for intraoperative wound management on the incidence of I-SSI following elective surgery for colorectal cancer.

Methods

Between November 2009 and February 2011, the data of 1,980 consecutive patients who underwent elective colorectal resection for colorectal cancer were prospectively collected from 19 affiliated hospitals. The incidence of and risk factors for I-SSI were investigated.

Results

Overall, 233 I-SSIs were identified (11.7 %). Forty-two possible risk factors were analyzed. Using a multivariate analysis, the independent risk factors for I-SSI were identified to be a high body mass index, previous laparotomy, chronic liver disease, wound length, contaminated wound class, creation or closure of an ostomy, right hemicolectomy procedure, the suture material used for fascial closure and the incidence of organ/space SSI.

Conclusion

To prevent I-SSI following elective colorectal surgery, it is crucial to avoid making large incisions and reduce fecal contamination whenever possible. A high quality randomized control trial is necessary to confirm the definitive intraoperative procedure(s) that can minimize the incidence of I-SSI.     

Anatomic Versus Nonanatomic Hepatectomy for a Solitary Hepatocellular Carcinoma

Background

It remains controversial whether anatomical resection (AR) improves the prognosis for hepatocellular carcinoma (HCC) or not. To our knowledge, there have been a few well-matched studies about this issue. The aim of the present study was to compare the recurrence-free survival of AR versus nonanatomical resection (NAR) for a solitary HCC using propensity score matching.

Methods

The present study included 236 patients who had a solitary HCC without macroscopic vessel thrombosis. Those patients were divided into AR (n?=?139) and NAR (n?=?97) groups. A propensity score matching was performed to minimize the effect of potential confounders.

Results

Sixty-four patients from each group were matched. Preoperative confounding factors were balanced between the two groups. The median recurrence-free survival times in the AR and NAR groups were 33.8 and 30.8 months, respectively (P?=?0.520). There were no significant differences in the intrahepatic recurrence pattern (P?=?0.097). Operative procedure was not a significant risk factor for recurrence in both uni- and multivariate analyses.

Conclusions

This case-matching study using a propensity score shows that there is no superiority of AR to NAR relevant to the recurrence-free survival in patients with a single HCC.     

Impact of concomitant use of proton pump inhibitors and clopidogrel or ticagrelor on clinical outcomes in patients with acute coronary syndrome

Background There is great debate on the possible adverse interaction between proton pump inhibitors (PPIs) and clopidogrel. In addition, whether the use of PPIs affects the clinical efficacy of ticagrelor remains less known. We aimed to determine the impact of concomitant administration of PPIs and clopidogrel or ticagrelor on clinical outcomes in patients with acute coronary syndrome (ACS) after percutaneous coronary intervention (PCI). Methods We retrospectively analyzed data from a “real world”, international, multi-center registry between 2003 and 2014 (n = 15,401) and assessed the impact of concomitant administration of PPIs and clopidogrel or ticagrelor on 1-year composite primary endpoint (all-cause death, re-infarction, or severe bleeding) in patients with ACS after PCI. Results Of 9,429 patients in the final cohort, 54.8% (n = 5165) was prescribed a PPI at discharge. Patients receiving a PPI were older, more often female, and were more likely to have comorbidities. No association was observed between PPI use and the primary endpoint for patients receiving clopidogrel (adjusted HR: 1.036; 95% CI: 0.903–1.189) or ticagrelor (adjusted HR: 2.320; 95% CI: 0.875–6.151) (P_interaction = 0.2004). Similarly, use of a PPI was not associated with increased risk of all-cause death, re-infarction, or a decreased risk of severe bleeding for patients treated with either clopidogrel or ticagrelor. Conclusions In patients with ACS following PCI, concomitant use of PPIs was not associated with increased risk of adverse outcomes in patients receiving either clopidogrel or ticagrelor. Our findings indicate it is reasonable to use a PPI in combination with clopidogrel or ticagrelor, especially in patients with a higher risk of gastrointestinal bleeding.