Development of Genome-Wide Intron Length Polymorphism (ILP) Markers in Tea Plant (Camellia sinensis) and Related Applications for Genetics Research.

The market value of tea is largely dependent on the tea species and cultivar. Therefore, it is important to develop efficient molecular markers covering the entire tea genome that can be used for the identification of tea varieties, marker-assisted breeding, and mapping important quantitative trait loci for beneficial traits. In this study, genome-wide molecular markers based on intron length polymorphism (ILP) were developed for tea trees. A total of 479, 1393, and 1342 tea ILP markers were identified using the PCR method in silico from the 'Shuchazao' scaffold genome, the chromosome-level genome of 'Longjing 43', and the ancient tea DASZ chromosome-level genome, respectively. A total of 230 tea ILP markers were used to amplify six tea tree species. Among these, 213 pairs of primers successfully characterize products in all six species, with 112 primer pairs exhibiting polymorphism. The polymorphism rate of primer pairs increased with the improvement in reference genome assembly quality level. The cross-species transferability analysis of 35 primer pairs of tea ILP markers showed an average amplification rate of 85.17% through 11 species in 6 families, with high transferability in Camellia reticulata and tobacco. We also used 40 pairs of tea ILP primers to evaluate the genetic diversity and population structure of C. tetracocca with 176 plants from Puan County, Guizhou Province, China. These genome-wide markers will be a valuable resource for genetic diversity analysis, marker-assisted breeding, and variety identification in tea, providing important information for the tea industry.

Study on the Drug Targets and Molecular Mechanisms of Rhizoma Curcumae in the Treatment of Nasopharyngeal Carcinoma Based on Network Pharmacology.

AIM: To analyse the target of Rhizoma Curcumae in nasopharyngeal carcinoma by using network pharmacological techniques and to explore the associated molecular mechanism. METHODS: The targets of nasopharyngeal carcinoma were retrieved from the GeneCards database. At the same time, the drug therapeutic targets of Rhizoma Curcumae were obtained from the TCMSP and SymMap databases. The data were imported into the STRING database and Cytoscape 3.7.1 to construct a network of "Chinese medicine component-target-disease" interactions; then, the intersection was screened as the core Rhizoma Curcumae antinasopharyngeal cancer targets. Through GO target function and KEGG pathway enrichment analyses of the core targets, we predicted the biological processes and key signalling pathways involved in the Rhizoma Curcumae treatment of nasopharyngeal carcinoma. RESULTS: Twenty-five core targets of Rhizoma Curcumae in nasopharyngeal carcinoma were mined: TP53, BCL2 ICAM1 RXRA, TLR3 and TLR9, TNF, PTGS2, IL-6, CTSD, MMP2, MMP9, MMP14, TIMP2, ABCC1, ABCB1, ABCG2, and so on. The results of visual analysis showed that the Rhizoma Curcumae treatment of nasopharyngeal carcinoma mainly involves leukocyte adhesion to vascular endothelial cells, positive regulation of NF-κB import into the nucleus, regulation of the reactive oxygen species biosynthetic and metabolic process, regulation of the chemokine biosynthetic and metabolic process, various cancer-related signalling pathways, and a variety of cytokine signal transduction pathways, such as the NF-κB, TLR, IL-17, and TNF signalling pathways. CONCLUSION: The core targets predicted by our research can be used as molecular markers for the treatment and prediction of nasopharyngeal carcinoma. The mechanism of Rhizoma Curcumae treatment in NPC may be related to immune regulatory pathways, the inhibition of cancer cell proliferation, metastasis, and angiogenesis, as well as the regulation of tumour microenvironment. Combined with the prediction of its associated mechanism of action, the core targets can provide targeted reference value for subsequent drug development related to Curcuma.

Caspase-3 activation in the guinea pig cochlea exposed to salicylate.

In the current study, we explored whether chronic salicylate exposure could induce apoptosis in outer hair cells (OHCs) and spiral ganglion neurons (SGNs) of the cochlea. Guinea pig received sodium salicylate (400 mg/kg/d) or saline vehicle for 10 consecutive days. Programmed cell death (PCD) executioner was evaluated with immunohistochemistry detection of activated caspase-3. Apoptosis was examined with a terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling (TUNEL) method. Repeated salicylate administration activated caspase-3 and caused apoptosis in OHCs and SGNs (p<0.01 vs. saline control for both measures and in both cell types). Cell counting showed a significant loss in OHCs (p<0.01 vs. saline control), but not in inner hair cells (IHCs). Transmission electron microscopy (TEM) revealed chromatin condensation and nucleus margination in salicylate-treated cochlea. Scanning electron microscopy (SEM) demonstrated stereociliary bundles breakdown and fusion at the apical of OHCs, villous matter was discovered to attach on the surface of SGNs. These findings suggest that long-term administration of high-dose salicylate can activate caspase-3 pathway to induce OHC and SGN apoptosis.

[Effects of sodium salicylate on the expressions of gamma-aminobutyricacid and glutamate and auditory response properties of the inferior colliculus neurons].

The effects of sodium salicylate (NaSA) on the expressions of gamma-aminobutyricacid (GABA) and glutamate (Glu), and auditory response properties of the inferior colliculus neurons in mice were studied. Thirty-six Kunming mice were divided into three groups: control group (saline injection); NaSA group (NaSA 450 mg/kg, i.p., each day for 15 d); NaSA + lidocaine group (NaSA 450 mg/kg + lidocaine 10 mg/kg, i.p., each day for 15 d). The expressions of GABA and Glu were examined with immunohistochemical method. The intensity-rate function, intensity-latency function and frequency-tuning curve were determined with extracellular electrophysiological recording. Results are as follows: (1) The expression of GABA in the NaSA and NaSA + lidocaine groups decreased remarkably compared with that in the control group; there was no noticeable difference between the NaSA and NaSA + lidocaine groups. The expression of Glu in the NaSA group increased significantly compared with that in the control and NaSA + lidocaine groups. No difference in the expression of Glu was found between the control and NaSA + lidocaine groups. (2) In NaSA group, the intensity-rate function displayed a non-monotonic pattern, rising at low intensity and descending at high intensity; the tip of frequency-tuning curves became broad after administration of NaSA. (3) The changes in intensity-rate function and intensity-latency function were not evident and the tips of the frequency-tuning curves sharpened in the NaSA + lidocaine group. These results suggest that administration of NaSA increases the expression of Glu-positive neurons and reduces that of GABA-positive neurons in the inferior colliculus. NaSA changes the auditory response properties of the inferior colliculus and lidocaine can reverse these changes.