Apheresis Induces Oxidative Stress in Blood Cells

Oxidative stress mediates damage to various cells and is thought to be involved in various pathologies, including hereditary and acquired hemolytic anemias. It is induced by a multitude of physiological and environmental factors, including extracorporeal manipulation of blood. As a result, hemodialysis induces oxidative damage to red blood cells, thereby increasing their susceptibility to hemolysis and shortening their life span. We studied the effect of apheresis on the oxidative status of blood components. Using flow cytometric measurements, we showed that red blood cells, lymphocytes, monocytes, and polymorphonuclear cells undergo oxidative stress induced by the procedure. Their reactive oxygen species and externalization of phosphatidylserine increased, while their levels of reduced glutathione decreased. This oxidative stress, which may be caused by a direct interaction with the membranous system, may lead to cellular abnormalities with clinical consequences such as hemolysis and platelet hyperactivation.     

Immune Intervention in Children with Type 1 Diabetes

Not only T cells but also B cells play a role in the autoimmune process. Both monoclonal antiCD3 and antiCD20 antibodies seem efficacious. However, such treatments need to be refined to minimize adverse events. Use of autoantigens to create tolerance is a concept with great potential. GAD65 treatment has shown efficacy without adverse events thus far, and administration of the insulin B chain shows interesting immunologic effects. Other more or less speculative approaches to modulate the immune process need further studies with good design. Risks that are too serious cannot be motivated. In addition, as the β cells may die even though the autoimmune process is stopped, protective measures may be valuable (eg, active insulin treatment, and perhaps interleukin-1 receptor antagonists to reduce the nonautoimmune inflammation). Combination of immune intervention, protection of the β cells, and stimulation of regeneration may lead to a milder disease or even a cure in the future, and prevention is no longer unrealistic.     

Thiol‐water proton exchange of glutathione,cysteine, and N‐acetylcysteine: Implications for CEST MRI

Amide‐, amine‐, and hydroxyl‐water proton exchange can generate MRI contrast through chemical exchange saturation transfer (CEST). In this study, we show that thiol‐water proton exchange can also generate quantifiable CEST effects under near‐physiological conditions (pH = 7.2 and 37°C) through the characterization of the pH dependence of thiol proton exchange in phosphate‐buffered solutions of glutathione, cysteine, and N‐acetylcysteine. The spontaneous, base‐catalyzed, and buffer‐catalyzed exchange contributions to the thiol exchange were analyzed. The thiol‐water proton exchange of glutathione and cysteine was found to be too fast to generate a CEST effect around neutral pH due to significant base catalysis. The thiol‐water proton exchange of N‐acetylcysteine was found to be much slower, yet still in the fast‐exchange regime with significant base and buffer catalysis, resulting in a 9.5% attenuation of the water signal at pH 7.2 in a slice‐selective CEST NMR experiment. Furthermore, the N‐acetylcysteine thiol CEST was also detectable in human serum albumin and agarose phantoms.     

Hepatitis C virus genotypes in two different patient cohorts in Johannesburg,South Africa

Previous studies in South African patients have shown that hepatitis C virus (HCV) prevalence is low (2.6%) and genotype 5 is the dominant genotype. This is the first sequence-based genotype study from this country on two different patient groups (haemophiliacs (H) and liver disease (LD) patients) over two time periods (2000–2002 and 2007). Genotype 5 was found to be the dominant genotype in the LD groups, but genotype 1 dominated in the H group. Genotype 2, and a higher prevalence of genotype 3a, was present in the H group when compared to previous South African studies. Genotypes 3 and 4 have been described here in the LD groups for the first time in South Africa, with subtypes 4c and 4g being identified. A separate cluster of genotype 5 sequences were found to have only one adenine at nucleotide position 107 of the 5′UTR, as previously reported. Subtyping in the less conserved NS5B region was used to further characterize the 2000–2002 isolates and validate 5′UTR typing. The study shows that by using reliable methods of genotyping, the prevalence and frequencies of HCV genotypes can be monitored in South Africa for better diagnosis and treatment of the disease.     

Antibacterial role for natural killer cells in host defense to Bacillus anthracis

Natural killer (NK) cells have innate antibacterial activity that could be targeted for clinical interventions for infectious disease caused by naturally occurring or weaponized bacterial pathogens. To determine a potential role for NK cells in immunity to Bacillus anthracis, we utilized primary human and murine NK cells, in vitro assays, and in vivo NK cell depletion in a murine model of inhalational anthrax. Our results demonstrate potent antibacterial activity by human NK cells against B. anthracis bacilli within infected autologous monocytes. Surprisingly, NK cells also mediate moderate antibacterial effects on extracellular vegetative bacilli but do not have activity against extracellular or intracellular spores. The immunosuppressive anthrax lethal toxin impairs NK gamma interferon (IFN-γ) expression, but neither lethal nor edema toxin significantly alters the viability or cytotoxic effector function of NK cells. Compared to human NK cells, murine NK cells have a similar, though less potent, activity against intracellular and extracellular B. anthracis. The in vivo depletion of murine NK cells does not alter animal survival following intranasal infection with B. anthracis spores in our studies but significantly increases the bacterial load in the blood of infected animals. Our studies demonstrate that NK cells participate in the innate immune response against B. anthracis and suggest that immune modulation to augment NK cell function in early stages of anthrax should be further explored in animal models as a clinical intervention strategy.     

Mechanical loading of stem cells for improvement of transplantation outcome in a model of acute myocardial infarction: the role of loading history

Stem cell therapy for tissue repair is a rapidly evolving field and the factors that dictate the physiological responsiveness of stem cells remain under intense investigation. In this study we hypothesized that the mechanical loading history of muscle-derived stem cells (MDSCs) would significantly impact MDSC survival, host tissue angiogenesis, and myocardial function after MDSC transplantation into acutely infarcted myocardium. Mice with acute myocardial infarction by permanent left coronary artery ligation were injected with either nonstimulated (NS) or mechanically stimulated (MS) MDSCs. Mechanical stimulation consisted of stretching the cells with equibiaxial stretch with a magnitude of 10% and frequency of 0.5?Hz. MS cell-transplanted hearts showed improved cardiac contractility, increased numbers of host CD31+ cells, and decreased fibrosis, in the peri-infarct region, compared to the hearts treated with NS MDSCs. MS MDSCs displayed higher vascular endothelial growth factor expression than NS cells in vitro. These findings highlight an important role for cyclic mechanical loading preconditioning of donor MDSCs in optimizing MDSC transplantation for myocardial repair.     

Microenvironments engineered by inkjet bioprinting spatially direct adult stem cells toward muscle- and bone-like subpopulations

In vivo, growth factors exist both as soluble and as solid-phase molecules, immobilized to cell surfaces and within the extracellular matrix. We used this rationale to develop more biologically relevant approaches to study stem cell behaviors. We engineered stem cell microenvironments using inkjet bioprinting technology to create spatially defined patterns of immobilized growth factors. Using this approach, we engineered cell fate toward the osteogenic lineage in register to printed patterns of bone morphogenetic protein (BMP) 2 contained within a population of primary muscle-derived stem cells (MDSCs) isolated from adult mice. This patterning approach was conducive to patterning the MDSCs into subpopulations of osteogenic or myogenic cells simultaneously on the same chip. When cells were cultured under myogenic conditions on BMP-2 patterns, cells on pattern differentiated toward the osteogenic lineage, whereas cells off pattern differentiated toward the myogenic lineage. Time-lapse microscopy was used to visualize the formation of multinucleated myotubes, and immunocytochemistry was used to demonstrate expression of myosin heavy chain (fast) in cells off BMP-2 pattern. This work provides proof-of-concept for engineering spatially controlled multilineage differentiation of stem cells using patterns of immobilized growth factors. This approach may be useful for understanding cell behaviors to immobilized biological patterns and could have potential applications for regenerative medicine.     

A Case Series of Robot-assisted Rectus Abdominis Flap Harvest for Pelvic Reconstruction: A Single Institution Experience

Study ObjectiveTo analyze outcomes and postoperative complications in patients undergoing robot-assisted rectus abdominis flap harvest for pelvic floor reconstruction.DesignCase series.SettingAcademic setting.PatientsPelvic reconstruction surgery patients.InterventionsThe rectus abdominis muscle flap can be used as a flap for pelvic reconstruction, providing a large volume of soft tissue that can be used in the treatment of many comorbid conditions, including genital fistulas, postradiation pelvic exenteration, and abdominoperineal resection defects. Intraperitoneal harvest of the rectus muscle using a robotic approach allows avoidance of laparotomy and subsequent disruption of the anterior rectus sheath, thus preserving the integrity of the abdominal wall.Measurements and Main ResultsA retrospective analysis of patient demographic and clinical characteristics was performed for all patients who underwent robot-assisted rectus abdominis flap harvest for pelvic floor reconstruction at our institution from October 1, 2016, to October 31, 2018. The postoperative complications analyzed included bowel obstructions, surgical site infections, emergency room visits, and need for readmission. Six patients (4 women and 2 men), with a mean age of 69.2 years (range = 57–79 years) and median follow-up time of 9.2 months (range = 5–12 months), were included. Muscle flap harvest was performed on the right side in 4 patients and on the left in 2 patients. The indications for reconstructive surgery included vesicovaginal fistula, complex pelvic organ prolapse, anterior and posterior exenteration, partial and total vaginectomy, partial vulvectomy, and abdominoperineal resection. Two patients received neoadjuvant chemoradiation. One of the 6 cases was converted to laparotomy; however, this was not owing to the rectus harvest. Three patients experienced no complications after reconstruction; 1 patient reported occasional abdominal pain; 1 patient had intermittent bowel obstruction; and 1 patient developed a pelvic abscess, requiring readmission. All 6 patients achieved satisfactory healing of the pelvic wound after robot-assisted rectus abdominis flap inset.ConclusionRobot-assisted rectus abdominis flap harvest for pelvic floor reconstruction is a reliable means of defect closure, despite the presence of substantial comorbidities and risk factors in this patient cohort. Patient selection and counseling are crucial to optimize surgical outcomes in this complex population.     

Discovery of a novel Mycobacterium tuberculosis lineage that is a major cause of tuberculosis in Rio de Janeiro, Brazil

The current study evaluated Mycobacterium tuberculosis isolates from Rio de Janeiro, Brazil, for genomic deletions. One locus in our panel of PCR targets failed to amplify in approximately 30% of strains. A single novel long sequence polymorphism (>26.3 kb) was characterized and designated RD(Rio). Homologous recombination between two similar protein-coding genes is proposed as the mechanism for deleting or modifying 10 genes, including two potentially immunogenic PPE proteins. The flanking regions of the RD(Rio) locus were identical in all strains bearing the deletion. Genetic testing by principal genetic group, spoligotyping, variable-number tandem repeats of mycobacterial interspersed repetitive units (MIRU-VNTR), and IS6110-based restriction fragment length polymorphism analysis cumulatively support the idea that RD(Rio) strains are derived from a common ancestor belonging solely to the Latin American-Mediterranean spoligotype family. The RD(Rio) lineage is therefore the predominant clade causing tuberculosis (TB) in Rio de Janeiro and, as indicated by genotypic clustering in MIRU-VNTR analysis, the most significant source of recent transmission. Limited retrospective reviews of bacteriological and patient records showed a lack of association with multidrug resistance or specific risk factors for TB. However, trends in the data did suggest that RD(Rio) strains may cause a form of TB with a distinct clinical presentation. Overall, the high prevalence of this genotype may be related to enhanced virulence, transmissibility, and/or specific adaptation to a Euro-Latin American host population. The identification of RD(Rio) strains outside of Brazil points to the ongoing intercontinental dissemination of this important genotype. Further studies are needed to determine the differential strain-specific features, pathobiology, and worldwide prevalence of RD(Rio) M. tuberculosis.     

Human-based evidence for the therapeutic potential of arginase inhibitors in cardiovascular diseases

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