Integrating Theoretical and Experimental Methods for Multi-Scale Tissue Engineering of the Annulus Fibrosus of the Intervertebral Disc

There is a critical need for tissue engineered replacements for diseased and degenerated intervertebral discs in order to assuage low back pain while restoring function to the spine. Despite progress by many research groups, it remains a challenge to engineer a replacement tissue that can withstand the complex, demanding loading environment of the spine. Due to the hierarchical organization of the intervertebral disc, successful recapitulation of its functional behavior requires replication of anatomic form and physiologic function over a wide range of length scales. In this work, the technology of electrospinning has been employed for tissue engineering of the annulus fibrosus (AF) of the intervertebral disc using a multi-scale approach. The mechanics of electrospun nanofibrous assemblies was first characterized, focusing on how microscopic organization translates to macroscopic mechanical function. Next, engineered tissues were formed by culturing cells on nanofibrous scaffolds, generating aligned, dense collagenous tissues that replicate the single lamellar organization of the AF. This technology was then expanded to engineer angle-ply laminates that replicated both the anatomic form and mechanical function of the native AF. Finally, these results were further extended to engineer an angle-ply fiber-reinforced hydrogel composite that parallels the macroscopic structural organization of the intervertebral disc. Throughout, mechanical testing and mathematical modeling was used to understand material behavior, quantify functional growth, and guide comparison between engineered AF constructs and native tissue benchmarks. Emphasis has been placed on reconciling compositional and structural observations with their macroscopic mechanical implications, utilizing theoretical models to understand these relationships, and using engineered tissues to improve our understanding of structure-function relations within native fiber-reinforced soft tissues

Chemokines in depression in health and in inflammatory illness: a systematic review and meta-analysis

Inflammatory illness is associated with depression. Preclinical work has shown that chemokines are linked with peripheral–central crosstalk and may be important in mediating depressive behaviours. We sought to establish what evidence exists that differences in blood or cerebrospinal fluid chemokine concentration discriminate between individuals with depression and those without. Following PRISMA guidelines, we systematically searched Embase, PsycINFO and Medline databases. We included participants with physical illness for subgroup analysis, and excluded participants with comorbid psychiatric diagnoses. Seventy-three studies met the inclusion criteria for the meta-analysis. Individuals with depression had higher levels of blood CXCL4 and CXCL7 and lower levels of blood CCL4. Sensitivity analysis of studies with only physically healthy participants identified higher blood levels of CCL2, CCL3, CCL11, CXCL7 and CXCL8 and lower blood levels of CCL4. All other chemokines examined did not reveal significant differences (blood CCL5, CCL7, CXCL9, CXCL10 and cerebrospinal fluid CXCL8 and CXCL10). Analysis of the clinical utility of the effect size of plasma CXCL8 in healthy individuals found a negative predictive value 93.5%, given the population prevalence of depression of 10%. Overall, our meta-analysis finds evidence linking abnormalities of blood chemokines with depression in humans. Furthermore, we have demonstrated the possibility of classifying individuals with depression based on their inflammatory biomarker profile. Future research should explore putative mechanisms underlying this association, attempt to replicate existing findings in larger populations and aim to develop new diagnostic and therapeutic strategies

Comparison of real-time PCR and hemagglutination assay for quantitation of human polyomavirus JC

Human polyomavirus JC (JCV), the etiological agent of the disease progressive multifocal leukoencephalopathy (PML) affects immunocompromised patients particularly patients with AIDS. In vitro studies of JCV infection are hampered by the lack of sensitive JCV quantitation tests. Although the hemagglutination (HA) assay has been routinely employed for in vitro quantitation of JCV, its sensitivity is severely limited. We have employed a real-time PCR assay which compares favorably with the HA assay for the in vitro quantitation of JCV. JCV(Mad1), propagated in primary human fetal glial (PHFG) cells in two independent laboratories, was purified and quantitated by the HA assay. Both batches of purified JCV(Mad1) were then serially diluted in Dulbecco's Modified Eagle's Medium to obtain HA titers ranging from 64 to 0.001 HA units (HAU) per 100 μL of virus suspension. DNA was extracted from 100 μL of virus suspension and eluted in 50 μL of buffer, and DNA amplification and quantitation were performed in the Bio-Rad iCycler iQ Multicolor Real-Time PCR Detection System using T-antigen as the target gene. Real-time PCR for quantitation of JCV was sensitive and consistently detected 1.8 × 10(1 )copies of JCV DNA, and as low as 0.001 HAU equivalent of JCV. Moreover, there was a strong linear correlation between the HA assay and the DNA copy number of JCV(Mad1). The intra-run and inter-run coefficients of variation for the JCV standard curve were 0.06% to 4.8% and 2.6% to 5.2%, respectively. Based on these data, real-time PCR can replace the less-sensitive HA assay for the reliable detection, quantitation and monitoring of in vitro JCV replication

Strategies to improve palatability and increase consumption intentions for Momordica charantia (bitter melon): A vegetable commonly used for diabetes management

<p>Abstract</p> <p>Background</p> <p>Although beneficial to health, dietary phytonutrients are bitter, acid and/or astringent in taste and therefore reduce consumer choice and acceptance during food selection. <it>Momordica charantia</it>, commonly known as bitter melon has been traditionally used in Ayurvedic and Chinese medicine to treat diabetes and its complications. The aim of this study was to develop bitter melon-containing recipes and test their palatability and acceptability in healthy individuals for future clinical studies.</p> <p>Methods</p> <p>A cross-sectional sensory evaluation of bitter melon-containing ethnic recipes was conducted among 50 healthy individuals. The primary endpoints assessed in this analysis were current consumption information and future intentions to consume bitter melon, before and after provision of attribute- and health-specific information. A convenience sample of 50, self-reported non-diabetic adults were recruited from the University of Hawaii. Sensory evaluations were compared using two-way ANOVA, while differences in stage of change (SOC) before and after receiving health information were analyzed by Chi-square (χ²) analyses.</p> <p>Results</p> <p>Our studies indicate that tomato-based recipes were acceptable to most of the participants and readily acceptable, as compared with recipes containing spices such as curry powder. Health information did not have a significant effect on willingness to consume bitter melon, but positively affected the classification of SOC.</p> <p>Conclusions</p> <p>This study suggests that incorporating bitter foods in commonly consumed food dishes can mask bitter taste of bitter melon. Furthermore, providing positive health information can elicit a change in the intent to consume bitter melon-containing dishes despite mixed palatability results.</p

Momordica charantia (bitter melon) inhibits primary human adipocyte differentiation by modulating adipogenic genes

<p>Abstract</p> <p>Background</p> <p>Escalating trends of obesity and associated type 2 diabetes (T2D) has prompted an increase in the use of alternative and complementary functional foods. <it>Momordica charantia </it>or bitter melon (BM) that is traditionally used to treat diabetes and complications has been demonstrated to alleviate hyperglycemia as well as reduce adiposity in rodents. However, its effects on human adipocytes remain unknown. The objective of our study was to investigate the effects of BM juice (BMJ) on lipid accumulation and adipocyte differentiation transcription factors in primary human differentiating preadipocytes and adipocytes.</p> <p>Methods</p> <p>Commercially available cryopreserved primary human preadipocytes were treated with and without BMJ during and after differentiation. Cytotoxicity, lipid accumulation, and adipogenic genes mRNA expression was measured by commercial enzymatic assay kits and semi-quantitative RT-PCR (RT-PCR).</p> <p>Results</p> <p>Preadipocytes treated with varying concentrations of BMJ during differentiation demonstrated significant reduction in lipid content with a concomitant reduction in mRNA expression of adipocyte transcription factors such as, peroxisome proliferator-associated receptor γ (PPARγ) and sterol regulatory element-binding protein 1c (SREBP-1c) and adipocytokine, resistin. Similarly, adipocytes treated with BMJ for 48 h demonstrated reduced lipid content, perilipin mRNA expression, and increased lipolysis as measured by the release of glycerol.</p> <p>Conclusion</p> <p>Our data suggests that BMJ is a potent inhibitor of lipogenesis and stimulator of lipolysis activity in human adipocytes. BMJ may therefore prove to be an effective complementary or alternative therapy to reduce adipogenesis in humans.</p

First record of saucer scallop Ylistrum balloti (Bernardi, 1861) from equatorial South China Sea

First record of saucer scallop Ylistrum balloti (Bernardi, 1861) from equatorial South China Sea. Ylistrum balloti is one of the Pectinidae species distributed within the Indo–Pacifc region. Recently, 15 live specimens of Y. balloti were recorded from the continental shelf of Sarawak, Malaysia. The main morphological characteristics were displayed on the outer valve, these being a brown–red colour and a clear concentric pattern of thin brown lines. The prominent internal ribbing numbers on both valves also helped species identifcation. The preliminary report of Y. balloti revealed that the species is present in Sarawak waters and can be further explored in the future

TLR7-mediated skin inflammation remotely triggers chemokine expression and leukocyte accumulation in the brain

Background: The relationship between the brain and the immune system has become increasingly topical as, although it is immune-specialised, the CNS is not free from the influences of the immune system. Recent data indicate that peripheral immune stimulation can significantly affect the CNS. But the mechanisms underpinning this relationship remain unclear. The standard approach to understanding this relationship has relied on systemic immune activation using bacterial components, finding that immune mediators, such as cytokines, can have a significant effect on brain function and behaviour. More rarely have studies used disease models that are representative of human disorders. Methods: Here we use a well-characterised animal model of psoriasis-like skin inflammation—imiquimod—to investigate the effects of tissue-specific peripheral inflammation on the brain. We used full genome array, flow cytometry analysis of immune cell infiltration, doublecortin staining for neural precursor cells and a behavioural read-out exploiting natural burrowing behaviour. Results: We found that a number of genes are upregulated in the brain following treatment, amongst which is a subset of inflammatory chemokines (CCL3, CCL5, CCL9, CXCL10, CXCL13, CXCL16 and CCR5). Strikingly, this model induced the infiltration of a number of immune cell subsets into the brain parenchyma, including T cells, NK cells and myeloid cells, along with a reduction in neurogenesis and a suppression of burrowing activity. Conclusions: These findings demonstrate that cutaneous, peripheral immune stimulation is associated with significant leukocyte infiltration into the brain and suggest that chemokines may be amongst the key mediators driving this response

Thermoluminescence of zircon: a kinetic model

The mineral zircon, ZrSiO4, belongs to a class of promising materials for geochronometry by means of thermoluminescence (TL) dating. The development of a reliable and reproducible method for TL dating with zircon requires detailed knowledge of the processes taking place during exposure to ionizing radiation, long-term storage, annealing at moderate temperatures and heating at a constant rate (TL measurements). To understand these processes one needs a kinetic model of TL. This paper is devoted to the construction of such amodel. The goal is to study the qualitative behaviour of the system and to determine the parameters and processes controlling TL phenomena of zircon. The model considers the following processes: (i) Filling of electron and hole traps at the excitation stage as a function of the dose rate and the dose for both (low dose rate) natural and (high dose rate) laboratory irradiation. (ii) Time dependence of TL fading in samples irradiated under laboratory conditions. (iii) Short time annealing at a given temperature. (iv) Heating of the irradiated sample to simulate TL experiments both after laboratory and natural irradiation. The input parameters of the model, such as the types and concentrations of the TL centres and the energy distributions of the hole and electron traps, were obtained by analysing the experimental data on fading of the TL-emission spectra of samples from different geological locations. Electron paramagnetic resonance (EPR) data were used to establish the nature of the TL centres. Glow curves and 3D TL emission spectra are simulated and compared with the experimental data on time-dependent TL fading. The saturation and annealing behaviour of filled trap concentrations has been considered in the framework of the proposed kinetic model and comparedwith the EPR data associated with the rare-earth ions Tb3+ and Dy3+, which play a crucial role as hole traps and recombination centres. Inaddition, the behaviour of some of the SiOmn− centres has been compared with simulation results.

Hantavirus in Northern Short-tailed Shrew, United States

Phylogenetic analyses, based on partial medium- and large-segment sequences, support an ancient evolutionary origin of a genetically distinct hantavirus detected by reverse transcription–PCR in tissues of northern short-tailed shrews (Blarina brevicauda) captured in Minnesota in August 1998. To our knowledge, this is the first evidence of hantaviruses harbored by shrews in the Americas

SANGER SEQUENCING TO DETERMINE THE ACCURACY OF BIOINFORMATIC SOFTWARE FOR CONFIRMING DROPOUT MUTATIONS IN THE SARS-COV-2 SPIKE GENE OBTAINED USING WHOLE GENOME SEQUENCING

Abstract of a poster to be presented at the 2022 JABSOM Biomedical SymposiumIntroduction: Whole genome sequencing (WGS) is a powerful tool that can be used to track SARS-CoV-2 variants and their spread through a population. New mutations that have led to the emergence of new variants have occurred in genes that encode important viral proteins such as the spike protein, resulting in dropout regions. Bioinformatic analysis can be used to predict these regions of dropout based on reference sequences, however the accuracy of these predictions are questionable. Therefore, the objective of this study is to conduct Sanger sequencing to determine the sequences of the dropout regions by using primers designed to target these regions. Methods: Nasal swabs collected from individuals confirmed to be SARS-CoV-2 PCR positive were obtained from various CLIA approved clinical laboratories across Oahu, Hawaii (UH IRB#21-07-820-21-1A). RNA extraction and RT-PCR using the ARTIC Network V3 primer pools were performed to amplify the whole genome of SARS-CoV-2. The purified PCR products were then processed for WGS at the Advanced Studies in Genomics, Proteomics and Bioinformatics (ASGPB) facility at the University of Hawaii at Manoa. The sequencing reads were mapped to the original Wuhan sequence (MN908947.3) and assembled into whole genomes using iVar workflow. To fill in the ambiguous bases, sequences from the GISAID database from the same lineage and similar collection dates were used as references. Spike gene consensus primers were designed to Sanger sequence the ARTIC primer pool binding sites which frequently contained the dropout. The final step is to compare the Sanger sequences with the predicted sequences based on the consensus reference sequences to determine the accuracy of the prediction. Results: Analysis of whole genome sequence reads showed that the region of the genome that was sequenced using primers 72 and 73 from the ARTIC primer pool frequently contained ambiguous bases, indicating dropouts in the region. ARTIC primers 72 and 73 bind to regions of the SARS-CoV-2 spike protein. Sanger sequencing is ongoing to confirm the dropout sequences. Discussion: WGS is used to track SARS-CoV-2 mutations that are integral to the development of diagnostics, therapeutics, and vaccines. Therefore, the accuracy of the sequences obtained using WGS is critical for patient care. Due to the high mutation rate of SARS-CoV-2, primers used in WGS can quickly become obsolete as the virus mutates and are unable to bind to highly variable regions of the genome, resulting in regions of dropout. Reference sequences can be used to fill in these ambiguous bases, however, this method is fallible. Sanger sequencing provides a reliable method to verify the accuracy of WGS when combined with bioinformatic predictions. Once validated, bioinformatic predictions can ultimately be used to reduce time and cost needed for efficient WGS. Acknowledgements: This research was supported by a COBRE grant (P30GM114737) from the Pacific Center for Emerging Infectious Diseases Research, a grant (P20GM103466) from the INBRE, National Institute of General Medical Sciences, and a grant (T37MD008636) from the National Institute on Minority Health and Health Disparities, NIH. We thank Dr. Jennifer Saito at the ASGPB, UH Manoa for her expertise with WGS, and Dr. Eileen Nakano and Dr. Sandra Chang for their assistance with sample procurement. We also thank the Tropical Medicine Clinical Laboratory, Kaiser Permanente Clinical Laboratory, and National Kidney Foundation of Hawaii for providing de-identified patient samples.COBRE grant (P30GM114737) from the Pacific Center for Emerging Infectious Diseases Research, a grant (P20GM103466) from the INBRE, National Institute of General Medical Sciences, and a grant (T37MD008636) from the National Institute on Minority Health and Health Disparities, NI