RESUMO
BACKGROUND: Cerebral ischemia-reperfusion injury (CI/RI) is a complex process leading to neuronal damage and death, with mitophagy implicated in its pathogenesis. However, the significance of mitophagy in CI/RI remains debated. HYPOTHESIS: We hypothesized that TRIM25 reduces ATAD3A expression by ubiquitinating ATAD3A, promoting mitophagy via the PINK1/Parkin pathway, and aggravating CI/RI. STUDY DESIGN: Rat middle cerebral artery occlusion (MCAO) followed by reperfusion and oxygen-glucose deprivation and reoxygenation (OGD/R) in PC12 cells were used as animal and cell models, respectively. METHODS: To evaluate the success of the CI/R modeling, TTC and HE staining were employed. The determination of serum biochemical indexes was carried out using relative assay kits. The Western Blot analysis was employed to assess the expression of ATAD3A, TRIM25, as well as mitophagy-related proteins (PINK1, Parkin, P62, and LC3II/LC3I). The mRNA levels were detected using QRT-PCR. Mitochondrial membrane potential was assessed through JC-1 staining. Mitosox Red Assay Kit was utilized to measure mitochondrial reactive oxygen species levels in PC12 cells. Additionally, characterization of the mitophagy structure was performed using transmission electron microscopy (TEM). RESULTS: Our findings showed down-regulation of ATAD3A and up-regulation of TRIM25 in both in vivo and in vitro CI/RI models. Various experimental techniques such as Western Blot, JC-1 staining, Mitosox assay, Immunofluorescence assay, and TEM observation supported the occurrence of PINK1/Parkin signaling pathway-mediated mitophagy in both models. ATAD3A suppressed mitophagy, while TRIM25 promoted it during CI/RI injury. Additionally, the results indicated that TRIM25 interacted with and ubiquitinated ATAD3A via the proteasome pathway, affecting ATAD3A protein stability and expression. CONCLUSION: TRIM25 promoted Pink1/Parkin-dependent excessive mitophagy by destabilizing ATAD3A, exacerbating CI/RI. Targeting TRIM25 and ATAD3A may offer therapeutic strategies for mitigating CI/RI and associated neurological damage.
RESUMO
rAj-Tspin, a soluble recombinant peptide from Apostichopus japonicus, can inhibit the integrin ß1 (ITGB1)/FAK/AKT signaling pathway in hepatocellular carcinoma (HCC) via cell epithelial-mesenchymal transition (EMT) and apoptosis. Zyxin (ZYX) is a focal adhesion protein that is considered a novel mediator of EMT and apoptosis. However, the inhibitory mechanisms of rAj-Tspin in HCC and whether it is related to ZYX are unclear. We examined the antitumor effect of rAj-Tspin on the Huh7 human HCC cell line and on a nude mouse model generated via subcutaneous injection or orthotopic intrahepatic transplantation of Huh7 cells. Our results revealed that rAj-Tspin strikingly reduced the viability and promoted the apoptosis of Huh7 cells and inhibited HCC tumor growth in nude mice. rAj-Tspin inhibited ITGB1 and ZYX protein expression in vivo and in vitro in a dose-dependent manner. Mechanistically, the FAK/AKT signaling pathway and the proliferation and invasion of HCC cells were suppressed upon ITGB1 and ZYX knockdown. Moreover, the effect of ITGB1 overexpression on the growth of HCC cells was inhibited by rAj-Tspin. In contrast, the promoting effect of ITGB1 overexpression could be inhibited by ZYX knockdown. ZYX knockdown had no effect on ITGB1 expression. These findings suggest that ZYX is required for the indispensable role of ITGB1 in rAj-Tspin-alleviated HCC and provide an important therapeutic target for HCC. In summary, the anti-HCC effect of rAj-Tspin potentially involves the regulation of the ITGB1/ZYX/FAK/AKT pathway, which in turn impacts EMT and apoptosis.
RESUMO
rLj-RGD3, a new member of the RGD (Arginine-Glycine-Aspartate)-motif toxin protein family obtained from Lampetra japonica by means of recombinant DNA techniques, has been demonstrated to be a platelet fibrinogen receptor antagonist and holds potential as a drug candidate for a specific indication. The present article reports an innovative validated highly sensitive and specific biotin-avidin enzyme linked immunosorbent assay (BA-ELISA) to provide a bio-analytical method for pharmacokinetic (PK) studies of rLj-RGD3. The concentration of picogram level rLj-RGD3 in rat plasma was measured using the developed double sandwich BA-ELISA assay, which used two mouse anti-rLj-RGD3 monoclonal antibodies that recognize different epitopes for capture and detection. This method was verified to be highly specific (blank plasma did not interfere with detection), precise (RSD <15%), and accurate (86%-113%). Absolute recovery was in the 94%-119% range. The calibration curve showed good linearity within the 50 to 1600 pg/mL range. The LOQ was as low as 50 pg/mL. The above validated assay was successfully employed to assess PK of rLj-RGD3 in rats. After i.v. and s.c. dosing with 30 µg/kg, the rLj-RGD3 plasma concentration declined bi-exponentially with time. This decay was best fitted to a two-compartment model. In conclusion, the BA-ELISA method described here meets all requirements for PK studies of rLj-RGD3 with an effective pharmacological dose in the µg/kg BW range.
Assuntos
Avidina , Biotina , Camundongos , Animais , Ratos , Bioensaio , Ensaio de Imunoadsorção Enzimática , Imunossupressores , Receptores de FibrinogênioRESUMO
Tail-welded blanks (TWBs) are widely used in automotive bodies to improve the structural performance and reduce weight. The stiffness and modal lightweight design optimisation of TWBs for automotive doors was performed in this study. The finite element model was validated through physical experiments. An L27 (312) Taguchi orthogonal array was used to collect the sample points. The multi-objective optimisation problem was transformed into a single-objective optimisation problem based on the grey relational degree. The optimal combination of structural design parameters was obtained for a tail-welded door using the proposed method, and the weight of the door structure was reduced by 2.83 kg. The proposed optimisation method has fewer iterations and a lower computational cost, enabling the design of lightweight TWBs.
RESUMO
The research shows that subjective feelings of people, such as emotions and fatigue, can be objectively reflected by electroencephalography (EEG) physiological signals Thus, an evaluation method based on EEG, which is used to explore auditory brain cognition laws, is introduced in this study. The brain cognition laws are summarized by analyzing the EEG power topographic map under the stimulation of three kinds of automobile sound, namely, quality of comfort, powerfulness, and acceleration. Then, the EEG features of the subjects are classified through a machine learning algorithm, by which the recognition of diversified automobile sound is realized. In addition, the Kalman smoothing and minimal redundancy maximal relevance (mRMR) algorithm is used to improve the recognition accuracy. The results show that there are differences in the neural characteristics of diversified automobile sound quality, with a positive correlation between EEG energy and sound intensity. Furthermore, by using the Kalman smoothing and mRMR algorithm, recognition accuracy is improved, and the amount of calculation is reduced. The novel idea and method to explore the cognitive laws of automobile sound quality from the field of brain-computer interface technology are provided in this study.
RESUMO
The toxic RGD peptide, rLj-RGD4, characterized by 4 (Arg-Gly-Asp) (RGD) motifs, is a novel mutant of rLj-RGD3 from the salivary gland of Lampetra japonica. Our previous study showd that rLj-RGD3 exerts a protective effect against cerebral ischemia injury in rats. Through the induction of middle cerebral artery occlusion/reperfusion (MCAO) injuries in rats, the present study investigated the effects and the mechanism through which rLj-RGD4 protects against ischemic stroke. In rats, treatment with rLj-RGD4 2 h after MCAO enhanced survival rate, improved movement and ameliorated the severity of brain infarction and apoptosis by diminishing pathological changes. This study demonstrated that rLj-RGD4 can reverse the downregulation of Bcl2, and the upregulation of Caspase-3. Mechanistic studies showed that rLj-RGD4 upregulated the expression levels of FAK, p-FAK, PI3K and p-Akt. In contrast, caspase-3 expression was inhibited. These results showed that rLj-RGD4 may reduce cerebral ischemia-reperfusion injury.