Objective:To explore the role of tropomyosin 3 (TPM3) in hypoxia/reoxygenation (H/R)-induced cardiomyocyte pyroptosis and fibroblast activation.Methods:Rat cardiomyocytes (H9c2 cells) were treated with H/R method to simulate myocardial ischemia/reperfusion (I/R) injury, and cell proliferation activity was evaluated with cell counting kit-8 (CCK8). The expression of TPM3 mRNA and protein was detected by quantitative real-time polymerase chain reaction (RT-qPCR) and Western blotting. H9c2 cells with stable TPM3-short hairpin RNA (shRNA) expression were constructed and treated with H/R (hypoxia for 3 hours, and reoxygenation for 4 hours). The expression of TPM3 was measured by RT-qPCR. The expressions of TPM3, pyroptosis-related proteins including caspase-1, NOD-like receptor protein 3 (NLRP3) and Gasdermin family proteins-N (GSDMD-N) were measured by Western blotting. The expression of caspase-1 was also observed by immunofluorescence assay. The levels of human interleukins (IL-1β, IL-18) in the supernatant were determined by enzyme-linked immunosorbent assay (ELISA) to elucidate the effect of sh-TPM3 on pyroptosis of cardiomyocytes. Rat myocardial fibroblasts were incubated with the above cell supernatant, and the expressions of human collagen Ⅰ, collagen Ⅲ, matrix metalloproteinase-2 (MMP-2), and matrix metalloproteinase inhibitor 2 (TIMP2) were detected by Western blotting to determine the effect of TPM3-interfered cardiomyocytes on the activation of fibroblasts under H/R conditions.Results:Compared with the control group, H/R treatment for 4 hours significantly decreased the survival rate of H9c2 cells [(25.81±1.90)% vs. (99.40±5.54)%, P < 0.01], promoted the expression of TPM3 mRNA and protein [TPM3/GAPDH (2 -ΔΔCt): 3.87±0.50 vs. 1, TPM3/β-Tubulin: 0.45±0.05 vs. 0.14±0.01, both P < 0.01], and promoted the expressions of caspase-1, NLRP3, GSDMD-N, and the enhanced release of cytokines IL-1β and IL-18 [cleaved caspase-1/caspase-1: 0.89±0.04 vs. 0.42±0.03, NLRP3/β-Tubulin: 0.39±0.03 vs. 0.13±0.02, GSDMD-N/β-Tubulin: 0.69±0.05 vs. 0.21±0.02, IL-1β (μg/L): 13.84±1.89 vs. 4.31±0.33, IL-18 (μg/L): 17.56±1.94 vs. 5.36±0.63, all P < 0.01]. However, compared with the H/R group, sh-TPM3 significantly weakened the promoting effects of H/R on these proteins and cytokines [cleaved caspase-1/caspase-1: 0.57±0.05 vs. 0.89±0.04, NLRP3/β-Tubulin: 0.25±0.04 vs. 0.39±0.03, GSDMD-N/β-Tubulin: 0.27±0.03 vs. 0.69±0.05, IL-1β (μg/L): 8.56±1.22 vs. 13.84±1.89, IL-18 (μg/L): 9.34±1.04 vs. 17.56±1.94, all P < 0.01]. In addition, the expressions of collagen Ⅰ, collagen Ⅲ, TIMP2, and MMP-2 in myocardial fibroblasts were significantly increased by the cultured supernatants from the H/R group (collagen Ⅰ/β-Tubulin: 0.62±0.05 vs. 0.09±0.01, collagen Ⅲ/β-tubulin: 0.44±0.03 vs. 0.08±0.00, TIMP2/β-tubulin: 0.73±0.04 vs. 0.20±0.03, TIMP2/β-Tubulin: 0.74±0.04 vs. 0.17±0.01, all P < 0.01). However, these boosting effects were weakened by sh-TPM3 (collagen Ⅰ/β-Tubulin: 0.18±0.01 vs. 0.62±0.05, collagen Ⅲ/β-Tubulin: 0.21±0.03 vs. 0.44±0.03, TIMP2/β-Tubulin: 0.37±0.03 vs. 0.73±0.04, TIMP2/β-Tubulin: 0.45±0.03 vs. 0.74±0.04, all P < 0.01). Conclusion:Interference with TPM3 can alleviate H/R-induced cardiomyocyte pyroptosis and fibroblast activation, suggesting that TPM3 may be a potential target of myocardial I/R injury.

Although phosphodiesterase type 5 inhibitors (PDE5Is) are a revolution in the treatment of erectile dysfunction (ED) and have been marketed since 1998, they cannot restore pathological changes in the penis. Low-energy shock wave therapy (LESWT) has been developed for treating ED, and clinical studies have shown that LESWT has the potential to affect PDE5I non-responders with ED with few adverse effects. Animal studies have shown that LESWT significantly improves penile hemodynamics and restores pathological changes in the penis of diabetic ED animal models. Although the mechanisms remain to be investigated, recent studies have reported that LESWT could partially restore corpus cavernosum fibromuscular pathological changes, endothelial dysfunction, and peripheral neuropathy. LESWT could be a novel modality for treating ED, and particularly PDE5I non-responders with organic ED, in the near future. However, further extensive evidence-based basic and clinical studies are needed. This review intends to summarize the scientific background underlying the effect of LESWT on ED.
Animals ; Erectile Dysfunction* ; Hemodynamics ; Lithotripsy ; Male ; Models, Animal ; Penis ; Peripheral Nervous System Diseases ; Phosphodiesterase 5 Inhibitors ; Shock*

Objective:To explore the pathogenic variants of a family with syndromic deafness by high-throughput sequencing.Methods:The family was from Puyang City, Henan Province, and had four members, including two with syndromic deafness. The proband and his sister had congenital deafness, and their parents had normal phenotypes. The clinical phenotype of the family was characterized using clinical examinations and pedigree analysis. The clinical examinations included imaging examination, audiometry (pure tone audiometry, acoustic immittance, brainstem auditory evoked potential, and otoacoustic emission), vestibular function test, and ophthalmic examination (visual acuity test, visual field test, fundus examination, visual evoked potential, and electroretinogram). Target exome sequencing of 129 known deafness genes and bioinformatics analysis were used to screen suspected pathogenic variants. Sanger sequencing and minigene assay were used to verify and functionally investigate the mutation detected, respectively. According to the standards and guidelines for interpreting genetic variants proposed by the American College of Medical Genetics and Genomics, the variants c.6049G>A and c.8699A>G were classified as pathogenic/likely pathogenic, and the variant c.9856C>G was classified as variants of uncertain significance.Results:The probands and his sister had severe sensorineural hearing loss with decreased binocular vision, night blindness, decreased peripheral visual field sensitivity and partial visual field defect, and normal vestibular function. Both of them had three CDH23 mutations, including CDH23 (NM_022124.5) c.6049G>A (p.Gly2017Ser),c.9856C>G (p.His3286Asp), and c.8699A>G (p. Asp2900Gly), The first two were inherited from the father, and the last one was from the mother. The missense variants c.9856C>G and c.8699A>G were not included in the gnomad database. The missense mutation c.6049G>A was located in the last position of exon 46 and was predicted to affect splicing by bioinformatics software. The minigene experiment showed that the mutation cause exon skipping of exon 46, resulting in an abnormal protein. Conclusions:Compound heterozygous variations of the CDH23 are the leading cause of USH1D in the family. This study confirms that the compound heterozygosity of splicing and missense variants of the CDH23 gene could lead to USH1D.