Methods: hTenowere isolated from human hamstring tendon. Presence of insulin receptor beta (INSR-β) on normal tendon tissues and the hTeno monolayer culture were analyzed by immunofluorescence staining. The presence of Glucose Transporter Type 1 (GLUT1) and Glucose Transporter Type 4 (GLUT4) on the hTeno monolayer culture were also analyzed by immunofluorescence staining. Primary hTeno were treated with 0.008, 0.08, 0.8 and 8.0 µM of TNF-α, with and without insulin supplement. Outcome measures include 2-[N-(7-nitrobenz-2-oxa-1,3-diazol-4-yl) amino]-2-deoxy-d-glucose (2-NBDG) assay to determine the glucose uptake activity; colourimetric total collagen assay to quantify the total collagen expression levels; COL-I ELISA assay to measure the COL-I expression levels and real-time qPCR to analyze the mRNA gene expressions levels of Scleraxis (SCX), Mohawk (MKX), type I collagen (COL1A1), type III collagen (COL3A1), matrix metalloproteinases (MMP)-9 and MMP-13 in hTeno when treated with TNF-α. Apoptosis assay for hTeno induced with TNF-α was conducted using Annexin-V FITC flow cytometry analysis.
Results: Immunofluorescence imaging showed the presence of INSR-β on the hTeno in the human Achilles tendon tissues and in the hTeno in monolayer culture. GLUT1 and GLUT4 were both positively expressed in the hTeno. TNF-α significantly reduced the insulin-mediated 2-NBDG uptake in all the tested concentrations, especially at 0.008 µM. Total collagen expression levels and COL-I expression levels in hTeno were also significantly reduced in hTeno treated with 0.008 µM of TNF-α. The SCX, MKX and COL1A1 mRNA expression levels were significantly downregulated in all TNF-α treated hTeno, whereas the COL3A1, MMP-9 and MMP-13 were significantly upregulated in the TNF-α treated cells. TNF-α progressively increased the apoptotic cells at 48 and 72 h.
Conclusion: At 0.008 µM of TNF-α, an IR condition was induced in hTeno, supported with the significant reduction in glucose uptake, as well as significantly reduced total collagen, specifically COL-I expression levels, downregulation of candidate tenogenic markers genes (SCX and MKX), and upregulation of ECM catabolic genes (MMP-9 and MMP-13). Development of novel IR model in hTeno provides an insight on how tendon homeostasis could be affected and can be used as a tool for further discovering the effects on downstream molecular pathways, as the implication for diabetic tendinopathy.
Methodology: The pangolin P. fungorum (pangolin Pf) genome has a genomic size of approximately 7.7 Mbps with N50 of 69,666 bps. Our study showed that pangolin Pf is a Paraburkholderia fungorum supported by evidence from the core genome SNP-based phylogenetic analysis and the ANI analysis. Functional analysis has shown that the presence of a considerably large number of genes related to stress response, virulence, disease, and defence. Interestingly, we identified different types of secretion systems in the genome of pangolin Pf, which are highly specialized and responsible for a bacterium's response to its environment and in physiological processes such as survival, adhesion, and adaptation. The pangolin Pf also shared some common virulence genes with the known pathogenic member of the Burkholderiales. These genes play important roles in adhesion, motility, and invasion.
Conclusion: This study may provide better insights into the functions, secretion systems and virulence of this pangolin-associated bacterial strain. The addition of this genome sequence is also important for future comparative analysis and functional work of P. fungorum.
Methods: We carried out fogging with Pyrethroid insecticide (Detral 2.5 EC) at 10 different sites in a forest situated in the state of Selangor, Peninsular Malaysia. Across the sites, we counted the numbers of knocked-down invertebrates and identified them based on morphology to different taxa. We constructed Bayesian hierarchical Poisson regression models to investigate the effects of fogging on: (1) a target invertebrate taxon (Diptera) 3-h post-fogging; (2) selected non-target invertebrate taxa 3-h post-fogging; and (3) an invertebrate pollinator taxon (Lepidoptera) 24-h post-fogging.
Results: A total of 1,874 invertebrates from 19 invertebrate orders were knocked down by the fogging treatment across the 10 sites. Furthermore, 72.7% of the invertebrates counted 3-h post-fogging was considered dead. Our regression models showed that given the data and prior information, the probability that fogging had a negative effect on invertebrate taxa 3-h post-fogging was 100%, with reductions to 11% of the pre-fogging count of live individuals for the target invertebrate taxon (Diptera), and between 5% and 58% of the pre-fogging count of live individuals for non-target invertebrate taxa. For the invertebrate pollinator, the probability that fogging had a negative effect 24-h post-fogging was also 100%, with reductions to 53% of the pre-fogging count of live individuals.
Discussion: Our Bayesian models unequivocally demonstrate that fogging has detrimental effects on one pollinator order and non-target invertebrate orders, especially taxa that have comparatively lower levels of chitinisation. While fogging is effective in killing the target order (Diptera), no mosquitos were found dead in our experiment. In order to maintain urban biodiversity, we recommend that health authorities and the private sector move away from persistent insecticide fogging and to explore alternative measures to control adult mosquito populations.