Sequencing‐based microsatellite instability testing using as few as six markers for high‐throughput clinical diagnostics

Microsatellite instability (MSI) testing of colorectal cancers (CRCs) is used to screen for Lynch syndrome (LS), a hereditary cancer‐predisposition, and can be used to predict response to immunotherapy. Here, we present a single‐molecule molecular inversion probe and sequencing‐based MSI assay and demonstrate its clinical validity according to existing guidelines. We amplified 24 microsatellites in multiplex and trained a classifier using 98 CRCs, which accommodates marker specific sensitivities to MSI. Sample classification achieved 100% concordance with the MSI Analysis System v1.2 (Promega) in three independent cohorts, totaling 220 CRCs. Backward–forward stepwise selection was used to identify a 6‐marker subset of equal accuracy to the 24‐marker panel. Assessment of assay detection limits showed that the 24‐marker panel is marginally more robust to sample variables than the 6‐marker subset, detecting as little as 3% high levels of MSI DNA in sample mixtures, and requiring a minimum of 10 template molecules to be sequenced per marker for >95% accuracy. BRAF c.1799 mutation analysis was also included to streamline LS testing, with all c.1799T>A variants being correctly identified. The assay, therefore, provides a cheap, robust, automatable, and scalable MSI test with internal quality controls, suitable for clinical cancer diagnostics.     

In Vitro Elution Characteristics of Vancomycin in a Composite Calcium Phosphate/Calcium Sulfate Bone Substitute

Background

Periprosthetic joint infection is a particularly difficult orthopedic problem, complicating a growing number of revision procedures. Joint debridement and systemic antibiotics are the mainstay of treatment, yet difficulty remains in maintaining a minimum inhibitory concentration of antibiotic at the localized site of infection.

Study Aims

This study analyzes the elution characteristics of a 40%bwt calcium phosphate–60%bwt calcium sulfate composite, at varying concentrations of vancomycin.

Methods

Four groups of varying concentrations of vancomycin (2.63%bwt, 5.13%, 9.76%, and 17.78%) were mixed with one pack of the composite cement. At designated time intervals up to 28 days, the antibiotic concentration was detected using fluorescence polarization immunoassay and the elution trends compared.

Results

The elution rate of each of the four groups decreased over time. At almost all of the intervals, the elution rates of the higher concentration groups were significantly higher than the lower concentration groups (P < 0.05).

Conclusions

Calcium sulfate reabsorbs over a prolonged period, producing porosity which allows for new bone ingrowth through occupation of osteoprogenitor cells and osteoblasts; while calcium phosphate acts as a long-term osteoconductive matrix.

Clinical Relevance

The results of this study suggest that vancomycin can be mixed affectively with a calcium sulfate/phosphate composite, both maintaining stability and eluting gradually over a clinically relevant period of time.     

The lack of correlation between radiographic findings and cartilage integrity

Total knee arthroplasty is a common treatment of osteoarthritis, although unicompartmental knee arthroplasties are frequently used to retain unaffected compartments. Joint space width (JSW) is a major factor in determining treatment. We examined the relationship between JSW and cartilage quality in 60 patients undergoing total knee arthroplasty to assess its accuracy in representing cartilage degradation. Radiographic JSW was recorded, whereas the unaffected compartment of each tibial plateau was examined postoperatively using Collins, Mankin, and Kellgren and Lawrence scores. No correlation was seen between visual or histologic grading and JSW. Histology more accurately represented cartilage quality, yet it is impractical to obtain preoperatively; thus, JSW is the main mode of assessment. However, using JSW solely to indicate unicompartmental knee arthroplasty may overlook disease in apparently unaffected compartments.     

Intraosseous ganglion cysts of the carpus: current practice

Intraosseous cysts of the carpal bones are an infrequent cause of chronic wrist pain. The main body of work has investigated their occurrence in the proximal carpus, with limited incidence in the distal row. We review the current literature on the treatment of symptomatic carpal cysts following the report of a 17-year-old male with a 12-month history of progressive right wrist pain due to an intraosseous ganglion of the trapezoid. This review explores the pathology of carpal cysts, their varying presentation and current treatments.     

D-ribose-L-cysteine enhances memory task,attenuates oxidative stress and acetyl-cholinesterase activity in scopolamine amnesic mice

d -Ribose-l -cysteine (DRLC) is an analogue of cysteine that has been shown to boost cellular antioxidant capacity by enhancing intracellular biosynthesis of glutathione (GSH). Deficiency of GSH has been implicated in the pathogenesis of Alzheimer's disease (AD), a neurodegenerative disorder associated with loss of memory. Thus, the use of antioxidants to prevent or retard the progression of memory deteriorations in persons with AD has been the focus of intense investigations. This study was carried out to evaluate the effects of DRLC on memory and scopolamine-induced amnesia, acetyl-cholinesterase activity, and oxidative stress in mice. Male Swiss mice were given oral administration of saline (10 ml/kg), DRLC (25, 50, and 100 mg/kg) or donepezil (1 mg/kg) 30 min before testing for memory performance using Y-maze and object recognition models. Another set of mice were also pretreated orally with saline, DRLC (25, 50, and 100 mg/kg) or donepezil (1 mg/kg) but in combination with scopolamine (3 mg/kg, i.p.) daily for 7 days. Thirty minutes after treatment on Day 7, memory function was then evaluated. The brain levels of acetyl-cholinesterase and oxidative stress parameters were assayed. DRLC significantly (p < .05) enhanced memory performance and attenuated scopolamine-induced amnesia. Increased acetyl-cholinesterase activity and oxidative stress, as shown by decreased antioxidant substrates (glutathione and catalase) and elevated malondialdehyde contents in mice with scopolamine amnesia were also attenuated by DRLC. Our findings suggest that inhibition of oxidative stress and acetyl-cholinesterase activity might contribute to the potential benefit of DRLC in persons with amnesia.     

“Their place is beyond the town's border”: A qualitative exploration of stigma associated with tuberculosis in rural and urban areas of Lagos,Nigeria

Tuberculosis (TB) remains a major public health challenge in Nigeria with a minimum yield of various TB control efforts due to sociocultural determinants of health including TB-associated stigma. Therefore, to achieve the Sustainable Development Goal targets for TB control, an understanding and reduction in TB-associated stigma is necessary. The study aims to explore the perspective of community members and investigate the possible ways of mitigating TB-associated stigma in rural and urban areas in Lagos State, Nigeria. Eight focus group discussions (FGD) were conducted among eight homogenous groups of participants living in the community in rural and urban areas of Lagos state who were stratified by gender, between July and November 2017. Analysis of data was done using the modified grounded theory. A total of 86 participants took part in the FGDs. There were various stigmatising behaviours towards people infected with TB in rural and urban communities studied. This includes: Not willing to eat with people suffering from TB, withdrawal from TB patients in social gatherings, verbal abuse of TB patients and refusing to visit their houses because of their illness. There were also misconceptions about the cause of TB in our study which includes spiritual attack, ingestion of cat hair and inhalation of dust. However, participants in the study believed that mitigating the effect of TB-associated stigma will require adequate community education on TB, provision of financial and emotional support to the patients, as well as the involvement of community leaders in TB control activities and stigma reduction interventions. TB-associated stigma exists in rural and urban communities, with a lack of appropriate knowledge of TB and fear of infection as a major determinant in rural and urban areas respectively. Health education and sensitisation about TB, with community leaders as champions could help to mitigate the effect of TB-associated stigma.