SARS-CoV-2 produces a microRNA CoV2-miR-O8 in patients with COVID-19 infection.

Many viruses produce microRNAs (miRNAs), termed viral miRNAs (v-miRNAs), with the capacity to target host gene expression. Bioinformatic and cell culture studies suggest that SARS-CoV-2 can also generate v-miRNAs. This patient-based study defines the SARS-CoV-2 encoded small RNAs present in nasopharyngeal swabs of patients with COVID-19 infection using small RNA-seq. A specific conserved sequence (CoV2-miR-O8) is defined that is not expressed in other coronaviruses but is preserved in all SARS-CoV-2 variants. CoV2-miR-O8 is highly represented in nasopharyngeal samples from patients with COVID-19 infection, is detected by RT-PCR assays in patients, has features consistent with Dicer and Drosha generation as well as interaction with Argonaute and targets specific human microRNAs.

Calcium food sources in primary school children with low calcium intakes in Kuala Lumpur.

Background: Adequate calcium intake at an early age is crucial to achieving peak bone mass. Nevertheless, low calcium intake is common in Malaysian children. Aim: This study examined the calcium food sources and factors associated with low calcium intake among 243 children aged 9-11 years in Kuala Lumpur. Methods: Diet histories and bone density were measured. Results: The mean calcium intake was 370 ± 187 mg/day. The main contributors to calcium intake were beverages (19.2%), cereal (18.6%), milk and dairy (13.0%), meat and poultry (12.9%), and fish and seafood (10.1%). Within each food group, calcium-contributing foods tend to be from low bioavailability sources such as rice, cocoa-based and malted drinks, and chicken rather than milk. Children who practised regular meals, ate breakfast and snacks and consumed milk more than one serving daily have a higher calcium intake. Conclusion: In conclusion, public health strategies to improve the status of low calcium intake and poor choices of calcium-rich foods are needed to optimise bone health in this population.

Porocarcinoma with orbital metastasis, a case report with review of literature.

We present a patient who presented with an orbital mass lesion which was a metastatic lesion from a porocarcinoma of the scalp with progressive deterioration of the patient.A 78-year-old male presented with functional decline and a rapidly growing scalp lesion of 3 months duration. In addition to the scalp lesion, Computed Tomography showed an incidental finding of a left lateral orbital wall tumour. Fine-needle aspiration of the two lesions revealed malignant cells with similar morphologies. Punch biopsy of the scalp lesion showed histological features suggestive of a porocarcinoma. Patient underwent palliative radiotherapy and immunotherapy and subsequently succumbed to the disease.

Parallel recovery of chromatin accessibility and gene expression dynamics from frozen human regulatory T cells.

Epigenetic features such as DNA accessibility dictate transcriptional regulation in a cell type- and cell state- specific manner, and mapping this in health vs. disease in clinically relevant material is opening the door to new mechanistic insights and new targets for therapy. Assay for Transposase Accessible Chromatin Sequencing (ATAC-seq) allows chromatin accessibility profiling from low cell input, making it tractable on rare cell populations, such as regulatory T (Treg) cells. However, little is known about the compatibility of the assay with cryopreserved rare cell populations. Here we demonstrate the robustness of an ATAC-seq protocol comparing primary Treg cells recovered from fresh or cryopreserved PBMC samples, in the steady state and in response to stimulation. We extend this method to explore the feasibility of conducting simultaneous quantitation of chromatin accessibility and transcriptome from a single aliquot of 50,000 cryopreserved Treg cells. Profiling of chromatin accessibility and gene expression in parallel within the same pool of cells controls for cellular heterogeneity and is particularly beneficial when constrained by limited input material. Overall, we observed a high correlation of accessibility patterns and transcription factor dynamics between fresh and cryopreserved samples. Furthermore, highly similar transcriptomic profiles were obtained from whole cells and from the supernatants recovered from ATAC-seq reactions. We highlight the feasibility of applying these techniques to profile the epigenomic landscape of cells recovered from cryopreservation biorepositories.

Effect of soluble corn fibre and calcium supplementation on bone mineral content and bone mineral density in preadolescent Malaysian children-a double-blind randomised controlled trial (PREBONE-Kids Study).

Soluble corn fibre (SCF) with calcium did not improve bone indices after 1 year in preadolescent children. INTRODUCTION: SCF has been reported to improve calcium absorption. We investigated the long-term effect of SCF and calcium on bone indices of healthy preadolescent children aged 9-11 years old. METHODS: In a double-blind, randomised, parallel arm study, 243 participants were randomised into four groups: placebo, 12-g SCF, 600-mg calcium lactate gluconate (Ca) and 12-g SCF + 600-mg calcium lactate gluconate (SCF + Ca). Total body bone mineral content (TBBMC) and total body bone mineral density (TBBMD) were measured using dual-energy X-ray absorptiometry at baseline, 6 and 12 months. RESULTS: At 6 months, SCF + Ca had a significant increase in TBBMC from baseline (27.14 ± 6.10 g, p = 0.001). At 12 months, there was a significant increase in TBBMC from baseline in the SCF + Ca (40.28 ± 9.03 g, p = 0.001) and SCF groups (27.34 ± 7.93 g, p = 0.037). At 6 months, the change in TBBMD in the SCF + Ca (0.019 ± 0.003 g/cm2) and Ca (0.014 ± 0.003 g/cm2) groups was significantly different (p < 0.05) from SCF (0.004 ± 0.002 g/cm2) and placebo (0.002 ± 0.003 g/cm2). However, the changes in TBBMD and TBBMC were not significantly different among groups at 12 months. CONCLUSION: SCF did not increase TBBMC and TBBMD in Malaysian children after 1 year although calcium supplementation increased TBBMD at 6 months. Further work is needed to fully understand the mechanism and health benefits of prebiotics in this study population. CLINICAL TRIAL REGISTRATION: https://clinicaltrials.gov/ct2/show/NCT03864172.

Eugeniifoline, a Pentacyclic Indole Alkaloid from Leuconotis eugeniifolia, and Configurational Revision of Synthetic Eugeniifoline Isolated from a Diversity-Enhanced Extract.

Eugeniifoline (1), a pentacyclic indole alkaloid with a five-membered ring E, was isolated for the first time as a natural product from the stem-bark extract of Leuconotis eugeniifolia. Eugeniifoline (1) was previously reported as a synthetic product from a diversity-enhanced extract, but with the configuration at C-21 reported as S (1a). The configuration at C-21 was revised to R as shown in 1, based on the NOE data, GIAO NMR calculations, and DP4+ probability analysis, as well as the TDDFT-ECD method.

All-in-focus fine needle aspiration biopsy imaging based on Fourier ptychographic microscopy.

Context: Cytology is the study of whole cells in diagnostic pathology. Unlike standard histologic thinly sliced specimens, cytologic preparations consist of preparations of whole cells where cells commonly cluster and aggregate. As such, cytology preparations are generally much thicker than histologic slides, resulting in large patches of defocus when examined under the microscope. A diagnostic aggregate of cells often cannot be viewed in focus together, requiring pathologists to continually manipulate the focal plane, complicating the task of accurately assessing the entire cellular aggregate and thus in making a diagnosis. Further, it is extremely difficult to acquire useful uniformly in-focus digital images of cytology preparations for applications such as remote diagnostic evaluations and artificial intelligence models. The predominant current method to address this issue is to acquire digital images at multiple focal planes of the entire slide, which demands long scanning time, complex and expensive scanning systems, and huge storage capacity. Aims: Here we report a unique imaging method that can acquire cytologic images efficiently and computationally render all-in-focus digital images that are highly compact. Methods and material: This method applies a metric-based digital refocusing to microscopy data collected with a Fourier ptychographic microscope (FPM). The digitally refocused patches of images are then synthesized into an all-in-focus image. Results: We report all-in-focus FPM results of thyroid fine needle aspiration (FNA) cytology samples, demonstrating our method's ability to overcome the height variance of 30 µm caused by cell aggregation, and rendering images at high resolution (corresponds to a standard microscope with objective NA of 0.75) and that are all-in-focus. Conclusions: This technology is applicable to standard microscopes, and we believe can have an impact on diagnostic accuracy as well as ease and speed of diagnosing challenging specimens. While we focus on cytology slides here, we anticipate this technology's advantages will translate well for histology applications. This technique also addresses the issue of remote rapid evaluation of cytology preparations. Finally, we believe that by resolving the focus heterogeneity issues in standard digital images, this technique is a critical advance for applying machine learning to cytology specimens.

What is the best prophylaxis against venous thromboembolism in Asians following total knee arthroplasty? A systematic review and network meta-analysis.

PURPOSE: Asians have a low venous thromboembolism (VTE) incidence following total knee arthroplasty (TKA). This systematic review and network meta-analysis was conducted to evaluate the best prophylaxis against VTE in Asians following total knee arthroplasty in current literature. MATERIALS AND METHODS: A systematic search of PubMed, Embase and CINAHL was conducted in adherence with the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA). Prophylaxis types were separated into low-molecular-weight heparin (LMWH), novel oral anti-coagulants (NOAC), mechanical-only prophylaxis (MOP) and no prophylaxis (NP). The primary outcome was VTE incidence, grouped according to diagnosis modality (ultrasound, venography, clinical). The secondary outcome was bleeding incidence, grouped into minor and major bleeding. RESULTS: Fourteen eligible articles, totalling 4259 patients, were pooled with the following significant results: NOACs had lower venography-diagnosed VTE incidence than LMWH (12.77%, p = 0.02) and NP (20.64, p < 0.001). MOP had lower venography-diagnosed VTE incidence than LMWH (23.72%, p < 0.001), NOACs (10.95%, p < 0.001) and NP (31.59%, p < 0.001) but, interestingly, a statistically higher ultrasound-diagnosed VTE incidence than LMWH (6.56%, p = 0.024) and NP (4.88%, p = 0.026). No significant differences were observed between prophylaxis types for symptomatic VTE, pulmonary embolism (PE) or death. LMWH and NOACs had a higher minor bleeding incidence than NP (11.71%, p < 0.001 and 6.33%, p < 0.02, respectively). No significant differences were observed between prophylaxis types for major bleeding incidence. CONCLUSION: NOACs are a superior form of chemoprophylaxis, compared with LMWH, in reducing venography-diagnosed VTE incidence with no added bleeding incidence. However, routine chemoprophylaxis may not be required as LMWH and NOACs do not appear to reduce symptomatic VTE incidence compared with MOP and NP with an increased minor bleeding incidence. Mechanical prophylaxis in the form of graduated compression stockings or intermittent pneumatic compression should be routinely considered with significantly lower rates of venography-diagnosed VTE compared with NP. On the basis of current evidence, we recommend an individualised approach to select the most appropriate prophylaxis type.

Lipidomic Analysis of Archival Pathology Specimens Identifies Altered Lipid Signatures in Ovarian Clear Cell Carcinoma.

Cancer metabolism is associated with the enhanced lipogenesis required for rapid growth and proliferation. However, the magnitude of dysregulation of diverse lipid species still requires significant characterization, particularly in ovarian clear cell carcinoma (OCCC). Here, we have implemented a robust sample preparation workflow together with targeted LC-MS/MS to identify the lipidomic changes in formalin-fixed paraffin-embedded specimens from OCCC compared to tumor-free ovarian tissue. We quantitated 340 lipid species, representing 28 lipid classes. We observed differential regulation of diverse lipid species belonging to several glycerophospholipid classes and trihexosylceramide. A number of unsaturated lipid species were increased in OCCC, whereas saturated lipid species showed a decrease in OCCC compared to the controls. We also carried out total fatty acid analysis and observed an increase in the levels of several unsaturated fatty acids with a concomitant increase in the index of stearoyl-CoA desaturase (SCD) in OCCC. We confirmed the upregulation of SCD (the rate-limiting enzyme for the synthesis of monounsaturated fatty acids) by immunohistochemistry (IHC) assays. Hence, by carrying out a mass spectrometry analysis of archival tissue samples, we were able to provide insights into lipidomic alterations in OCCC.

Factors associated with bone health status of Malaysian pre-adolescent children in the PREBONE-Kids Study.

BACKGROUND: Modifiable lifestyle factors and body composition can affect the attainment of peak bone mass during childhood. This study performed a cross-sectional analysis of the determinants of bone health among pre-adolescent (N = 243) Malaysian children with habitually low calcium intakes and vitamin D status in Kuala Lumpur (PREBONE-Kids Study). METHODS: Body composition, bone mineral density (BMD), and bone mineral content (BMC) at the lumbar spine (LS) and total body (TB) were assessed using dual-energy X-ray absorptiometry (DXA). Calcium intake was assessed using 1-week diet history, MET (metabolic equivalent of task) score using cPAQ physical activity questionnaire, and serum 25(OH) vitamin D using LC-MS/MS. RESULTS: The mean calcium intake was 349 ± 180 mg/day and mean serum 25(OH)D level was 43.9 ± 14.5 nmol/L. In boys, lean mass (LM) was a significant predictor of LSBMC (ß = 0.539, p < 0.001), LSBMD (ß = 0.607, p < 0.001), TBBMC (ß = 0.675, p < 0.001) and TBBMD (ß = 0.481, p < 0.01). Height was a significant predictor of LSBMC (ß = 0.346, p < 0.001) and TBBMC (ß = 0.282, p < 0.001) while fat mass (FM) (ß = 0.261, p = 0.034) and physical activity measured as MET scores (ß = 0.163, p = 0.026) were significant predictors of TBBMD in boys. Among girls, LM was also a significant predictor of LSBMC (ß = 0.620, p < 0.001), LSBMD (ß = 0.700, p < 0.001), TBBMC (ß = 0.542, p < 0.001) and TBBMD (ß = 0.747, p < 0.001). Calcium intake was a significant predictor of LSBMC (ß = 0.102, p = 0.034), TBBMC (ß = 0.122, p < 0.001) and TBBMD (ß = 0.196, p = 0.002) in girls. CONCLUSIONS: LM was the major determinant of BMC and BMD among pre-adolescent Malaysian children alongside other modifiable lifestyle factors such as physical activity and calcium intake.

Monomeric, Dimeric, and Trimeric Tropane Alkaloids from Pellacalyx saccardianus.

Seven new tropane alkaloids, including five monomeric (1-5), one dimeric (6), and one trimeric (7) 3α-nortropane ester, along with two known monomeric nortropane alkaloids (8 and 9), were isolated from the leaves and bark of Pellacalyx saccardianus. Their structures, including the absolute configuration of the enantiomeric pair of (±)-6, were elucidated by comprehensive spectroscopic analyses. Alkaloids 6 and 7 showed cytotoxicity toward human pancreatic cancer cell lines (AsPC-1, BxPC3, PANC-1, and SW1990). Alkaloids 1, 4, and 9 induced a smooth muscle relaxation effect comparable to that of atropine (Emax 106.1 ± 7.5%, 97.0 ± 5.2%, 100.9 ± 1.4%, 111.7 ± 1.7%, respectively) on isolated rat tracheal rings.

COVID-19 post-vaccination lymphadenopathy: Report of cytological findings from fine needle aspiration biopsy.

The coronavirus COVID-19 pandemic has spurred the rapid development of vaccines, with vaccination programmes already underway in many countries. Regional lymphadenopathy is one of the documented side effects of vaccination. We document the fine needle aspiration cytological findings of an enlarged supraclavicular lymph node in a 34-year-old Asian female following the first dose of the Pfizer-BioNTech COVID-19 mRNA vaccine, which appears to be the first such report in a premorbidly well patient with no known history of malignancy. The cytological findings featured a reactive pattern in keeping with follicular hyperplasia, with prominent germinal centre elements including lymphohistiocytic aggregates and tingible-body macrophages. Despite an increased proportion of larger lymphocytes, the overall pattern was in keeping with a reactive pattern, bearing in mind the temporal and geographic relation to the vaccination injection. In instances of localised lymphadenopathy, particularly in supraclavicular or axillary locations, pathologists should be cognizant of the possibility of post-vaccination reactive lymphadenopathy, and seek clinical and radiological hints favouring a benign process, whilst recognising potential morphological overlaps with lymphoproliferative disorders. Awareness of this diagnostic pitfall is especially important as COVID-19 vaccination coverage is ramped up worldwide, leading to an expected increase in incidence of post-vaccination reactive lymphadenopathy.

Design and strategies used for recruitment and retention in a double blind randomized controlled trial investigating the effects of soluble corn fiber on bone indices in pre-adolescent children (PREBONE-Kids study) in Malaysia.

BACKGROUND: Recruitment and retention in longitudinal nutrition intervention studies among children is challenging and scarcely reported. This paper describes the strategies and lessons learned from a 1-year randomized double-blind placebo-controlled trial among pre-adolescent children on the effects of soluble corn fiber (SCF) on bone indices (PREBONE-Kids). METHODS: Participants (9-11 years old) were recruited and randomized into 4 treatment groups (600 mg calcium, 12 g SCF, 12 g SCF plus 600 mg calcium and placebo). Interventions were consumed as a fruit-flavored powdered drink for 1-year. School-based recruitment was effective due to support on study benefits from parents and teachers, peer influence and a 2-weeks study run-in for participants to assess their readiness to commit to the study protocol. Retention strategies focused on building rapport through school-based fun activities, WhatsApp messaging, providing health screening and travel reimbursements for study measurements. Compliance was enhanced by providing direct on-site school feeding and monthly non-cash rewards. Choice of 2 flavors for the intervention drinks were provided to overcome taste fatigue. Satisfaction level on the manner in which the study was conducted was obtained from a voluntary sub-set of participants. RESULTS: The study successfully enrolled 243 participants within 6 months and retained 82.7% of the participants at the end of 1 year, yielding a drop-out rate of 17.3%. Compliance to the intervention drink was 85% at the start and remained at 78.7% at the end of 1 year. More than 95% of the participants provided good feedback on intervention drinks, rapport building activities, communication and overall study conduct. CONCLUSION: Successful strategies focused on study benefits, rapport building, frequent communication using social media and non-cash incentives helped improved compliance and retention rate. The lessons learned to maintain a high retention and compliance rate in this study provide valuable insights for future studies in a similar population.

Vitamin D Status Is Associated with Modifiable Lifestyle Factors in Pre-Adolescent Children Living in Urban Kuala Lumpur, Malaysia.

Studies on vitamin D status and its determinants in growing children in countries with ample sunshine such as Malaysia have been limited. The aim of our study was to determine factors associated with serum 25(OH)D concentrations such as lifestyle, dietary intake, anthropometry, and body composition in 243 pre-adolescent Malaysian children from low-income families living in Kuala Lumpur. This cross-sectional study measured bone density and body composition using dual-energy X-ray absorptiometry (DXA), while serum 25(OH)D was measured using LC-MS/MS. Time spent outdoors, body surface area exposed to sunlight, dietary intake, and physical activity level were assessed using questionnaires. Multiple linear regression and stepwise analysis were performed to identify significant predictors for serum 25(OH)D. About 69.4% had 25(OH)D < 50 nmol/L, and 18.9% were vitamin-D-deficient with 25(OH)D < 30 nmol/L. Girls had a nine-fold higher prevalence of vitamin D deficiency than boys. Body surface area exposed to sunlight, Sun Index, and fat mass were significant predictors of 25(OH)D concentrations in this population. Modifiable lifestyle factors such as sun exposure and reducing obesity are important public health guidance to ensure optimal vitamin D status in these children.