Validated numerical unrestrained occupant-seat crash scenarios for high-speed trains integrating experimental, computational, and inverse methods.

OBJECTIVE: Occupant impact safety is critical for train development. This paper proposes a systematic procedure for developing validated numerical occupant crash scenarios for high-speed trains by integrating experimental, computational, and inverse methods. METHODS: As the train interior is the most potentially injury-causing factor, the material properties were acquired by mechanical tests, and constitutive models were calibrated using inverse methods. The validity of the seat material constitutive model was further verified via drop tower tests. Finite element (FE) and multibody (MB) models of train occupant-seat interactions in frontal impact were established in LS-DYNA and MADYMO software, respectively, using the experimentally acquired materials/mechanical characteristics. Three dummy sled crash tests with different folding table and backrest configurations were conducted to validate the numerical occupant-seat models and to further assess occupant injury in train collisions. The occupant impact responses between dummy tests and simulations were quantitatively compared using a correlation and analysis (CORA) objective rating method. RESULTS: Results indicated that the experimentally calibrated numerical seat-occupant models could effectively reproduce the occupant responses in bullet train collisions (CORA scores >80%). Compared with the train seat-occupant MB model, the FE model could simulate the head acceleration with slightly more acceptable fidelity, however, the FE model CORA scores were slightly less than for the MB models. The maximum head acceleration was 30 g but the maximum HIC score was 17.4. When opening the folding table, the occupant's chest injury was not obvious, but the neck-table contact and "chokehold" may potentially be severe and require further assessment. CONCLUSIONS: This study demonstrates the value of experimental data for occupant-seat model interactions in train collisions and provides practical help for train interior safety design and formulation of standards for rolling stock interior passive safety.

TLR8 aggravates skin inflammation and fibrosis by activating skin fibroblasts in systemic sclerosis.

OBJECTIVES: Innate immunity significantly contributes to systemic sclerosis (SSc) pathogenesis. TLR8 is an important innate immune mediator that is implicated in autoimmunity and fibrosis. However, the expression, mechanism of action, and pathogenic role of TLR8 in SSc remain unclear. The aim of this study was to explore the roles and underlying mechanisms of TLR8 in SSc. METHODS: The expression of TLR8 was analyzed based on a public dataset and then verified in skin tissues and skin fibroblasts of SSc patients. The role of TLR8 in inflammation and fibrosis was investigated using a TLR8-overexpression vector, activator (VTX-2337), inhibitor (cu-cpt-8m), and TLR8 siRNA in skin fibroblasts. The pathogenic role of TLR8 in skin inflammation and fibrosis was further validated in a bleomycin (BLM)-induced mouse skin inflammation and fibrosis model. RESULTS: TLR8 levels were significantly elevated in SSc skin tissues and myofibroblasts, along with significant activation of the TLR8 pathway. In vitro studies showed that overexpression or activation of TLR8 by a recombinant plasmid or VTX-2337 upregulated IL-6, IL-1ß, COL I, COL III, and α-SMA in skin fibroblasts. Consistently, both TLR8-siRNA and cu-cpt-8m reversed the phenotypes observed in TLR8-activating fibroblasts. Mechanistically, TLR8 induces skin fibrosis and inflammation in a manner dependent on the MAPK, NF-κB, and SMAD2/3 pathways. Subcutaneous injection of cu-cpt-8m significantly alleviated BLM-induced skin inflammation and fibrosis in vivo. CONCLUSION: TLR8 might be a promising therapeutic target to improve the treatment strategy for SSc skin inflammation and fibrosis.

Thickened Retinal Nerve Fiber Layers Associated With High-Altitude Headache.

Purpose: This study aimed to quantify the different quadrants of the optic nerve head (ONH) and macular parameters and their changes during exposure to high altitude, and to assess their correlation with high-altitude headache (HAH). Methods: Spectral-domain optical coherence tomography (OCT) was used to quantify changes in the retinal structure in 109 healthy subjects during acute exposure to high altitude (3,700 m). Self-reported symptoms of HAH and acute mountain sickness AMS were assessed using Lake Louise Score (LLS), alongside measurements of physiological parameters (oxygen saturation [SpO2], heart rate [HR], hemoglobin level [Hb], and red blood cell [RBC] count). Measurements were taken before and after exposure to the high-altitude environment. The correlations of these parameters and changes at ONH were examined. Results: With the exposure to high altitude, the incidence of AMS was 44.0% and the frequency of HAH was 67.0% (54.1% mild, 12.9% moderate-severe). As for systemic parameters measured at high altitude, the participants exhibited significantly lower SpO2, higher resting HR, higher Hb, and a higher RBC (all p < 0.05). Key stereometric parameters used to describe ONH [superior, inferior, nasal, temporal, and mean retinal nerve fiber layer (RNFL) thickness] and macula (macular thickness) increased at high altitude compared with baseline. Most parameters of ONH changed, especially superior, inferior, and mean RNFL thickness (p < 0.05). There was a significant correlation between the ratios of RNFL at ONH and HAH [mean thickness (r = 0.246, p = 0.01); inferior (r = 0.216, p = 0.02); nasal (r = 0.193, p = 0.04)]. No associations between parameters of ONH and AMS or LLS were observed. Conclusion: The high-altitude environment can increase RNFL thickness at ONH. Furthermore, we found that the ratios of mean thickness, inferior area, and nasal area correlated positively with HAH, which provides new insights for understanding of the underlying pathological mechanisms of high-altitude retinopathy (HAR).

LDLR dysfunction induces LDL accumulation and promotes pulmonary fibrosis.

Treatments for pulmonary fibrosis (PF) are ineffective because its molecular pathogenesis and therapeutic targets are unclear. Here, we show that the expression of low-density lipoprotein receptor (LDLR) was significantly decreased in alveolar type II (ATII) and fibroblast cells, whereas it was increased in endothelial cells from systemic sclerosis-related PF (SSc-PF) patients and idiopathic PF (IPF) patients compared with healthy controls. However, the plasma levels of low-density lipoprotein (LDL) increased in SSc-PF and IPF patients. The disrupted LDL-LDLR metabolism was also observed in four mouse PF models. Upon bleomycin (BLM) treatment, Ldlr-deficient (Ldlr-/-) mice exhibited remarkably higher LDL levels, abundant apoptosis, increased fibroblast-like endothelial and ATII cells and significantly earlier and more severe fibrotic response compared to wild-type mice. In vitro experiments revealed that apoptosis and TGF-ß1 production were induced by LDL, while fibroblast-like cell accumulation and ET-1 expression were induced by LDLR knockdown. Treatment of fibroblasts with LDL or culture medium derived from LDL-pretreated endothelial or epithelial cells led to obvious fibrotic responses in vitro. Similar results were observed after LDLR knockdown operation. These results suggest that disturbed LDL-LDLR metabolism contributes in various ways to the malfunction of endothelial and epithelial cells, and fibroblasts during pulmonary fibrogenesis. In addition, pharmacological restoration of LDLR levels by using a combination of atorvastatin and alirocumab inhibited BLM-induced LDL elevation, apoptosis, fibroblast-like cell accumulation and mitigated PF in mice. Therefore, LDL-LDLR may serve as an important mediator in PF, and LDLR enhancing strategies may have beneficial effects on PF.

A Mitochondrial DNA Variant Elevates the Risk of Gallstone Disease by Altering Mitochondrial Function.

BACKGROUND AND AIMS: Gallstone disease (cholelithiasis) is a cholesterol-related metabolic disorders with strong familial predisposition. Mitochondrial DNA (mtDNA) variants accumulated during human evolution are associated with some metabolic disorders related to modified mitochondrial function. The mechanistic links between mtDNA variants and gallstone formation need further exploration. METHODS: In this study, we explored the possible associations of mtDNA variants with gallstone disease by comparing 104 probands and 300 controls in a Chinese population. We constructed corresponding cybrids using trans-mitochondrial technology to investigate the underlying mechanisms of these associations. Mitochondrial respiratory chain complex activity and function and cholesterol metabolism were assessed in the trans-mitochondrial cell models. RESULTS: Here, we found a significant association of mtDNA 827A>G with an increased risk of familial gallstone disease in a Chinese population (odds ratio [OR]: 4.5, 95% confidence interval [CI]: 2.1-9.4, P=1.2×10-4). Compared with 827A cybrids (haplogroups B4a and B4c), 827G cybrids (haplogroups B4b and B4d) had impaired mitochondrial respiratory chain complex activity and function and activated JNK and AMPK signaling pathways. Additionally, the 827G cybrids showed disturbances in cholesterol transport and accelerated development of gallstones. Specifically, cholesterol transport through the transporter ABCG5/8 was increased via activation of the AMPK signaling pathway in 827G cybrids. CONCLUSIONS: Our findings reveal that mtDNA 827A>G induces aberrant mitochondrial function and abnormal cholesterol transport, resulting in increased occurrence of gallstones. The results provide an important biological basis for the clinical diagnosis and prevention of gallstone disease in the future.

Mitochondrial DNA Haplogroup M7 Confers Disability in a Chinese Aging Population.

Mitochondrial DNA (mtDNA) haplogroups have been associated with functional impairments (i.e., decreased gait speed and grip strength, frailty), which are risk factors of disability. However, the association between mtDNA haplogroups and ADL disability is still unclear. In this study, we conducted an investigation of 25 mtSNPs defining 17 major mtDNA haplogroups for ADL disability in an aging Chinese population. We found that mtDNA haplogroup M7 was associated with an increased risk of disability (OR = 3.18 [95% CI = 1.29-7.83], P = 0.012). The survival rate of the M7 haplogroup group (6.1%) was lower than that of the non-M7 haplogroup group (9.5%) after a 6-year follow-up. In cellular studies, cytoplasmic hybrid (cybrid) cells with the M7 haplogroup showed distinct mitochondrial functions from the M8 haplogroup. Specifically, the respiratory chain complex capacity was significantly lower in M7 haplogroup cybrids than in M8 haplogroup cybrids. Furthermore, an obvious decreased mitochondrial membrane potential and 40% reduced ATP-linked oxygen consumption were found in M7 haplogroup cybrids compared to M8 haplogroup cybrids. Notably, M7 haplogroup cybrids generated more reactive oxygen species (ROS) than M8 haplogroup cybrids. Therefore, the M7 haplogroup may contribute to the risk of disability via altering mitochondrial function to some extent, leading to decreased oxygen consumption, but increased ROS production, which may activate mitochondrial retrograde signaling pathways to impair cellular and tissue function.

Contribution of Mitochondrial DNA Variation to Chronic Disease in East Asian Populations.

Mitochondria are the main producers of energy in eukaryotic cells. Mitochondrial dysfunction is associated with specific mitochondrial DNA (mtDNA) variations (haplogroups), and these variations can contribute to human disease. East Asian populations show enrichment of many mitochondrial haplogroups, including A, B, D, G, M7, M8, M9, N9, R9, and exhibit half of the known haplogroups of worldwide. In this review, we summarize the current research in the field of mtDNA variation and associated disease in East Asian populations and discuss the physiological and pathological relevance of mitochondrial biology. mtDNA haplogroups are associated with various metabolic disorders ascribed to altered oxidative phosphorylation. The same mitochondrial haplogroup can show either a negative or positive association with different diseases. Mitochondrial dynamics, mitophagy, and mitochondrial oxidative stress, ultimately influence susceptibility to various diseases. In addition, mitochondrial retrograde signaling pathways may have profound effects on nuclear-mitochondrial interactions, affecting cellular morphology, and function. Other complex networks including proteostasis, mitochondrial unfolded protein response and reactive oxygen species signaling may also play pivotal roles in metabolic performance.

Cell Type-Specific Modulation of Respiratory Chain Supercomplex Organization.

Respiratory chain complexes are organized into large supercomplexes among which supercomplex In + IIIn + IVn is the only one that can directly transfer electrons from NADH to oxygen. Recently, it was reported that the formation of supercomplex In + IIIn + IVn in mice largely depends on their genetic background. However, in this study, we showed that the composition of supercomplex In + IIIn + IVn is well conserved in various mouse and human cell lines. Strikingly, we found that a minimal supercomplex In + IIIn, termed "lowest supercomplex" (LSC) in this study because of its migration at the lowest position close to complex V dimers in blue native polyacrylamide gel electrophoresis, was associated with complex IV to form a supercomplex In + IIIn + IVn in some, but not all of the human and mouse cells. In addition, we observed that the 3697G>A mutation in mitochondrial-encoded NADH dehydrogenase 1 (ND1) in one patient with Leigh's disease specifically affected the assembly of supercomplex In + IIIn + IVn containing LSC, leading to decreased cellular respiration and ATP generation. In conclusion, we showed the existence of LSC In + IIIn + IVn and impairment of this supercomplex causes disease.

Exercise intolerance and developmental delay associated with a novel mitochondrial ND5 mutation.

The aim of this study was to evaluate the contribution of mitochondrial DNA (mtDNA) mutations in oxidative phosphorylation (OXPHOS) deficiency. The complete mitochondrial genomes of 41 families with OXPHOS deficiency were screened for mutations. Mitochondrial functional analysis was then performed in primary and cybrid cells containing candidate mutations identified during the screening. A novel mitochondrial NADH dehydrogenase 5 (ND5) m.12955A > G mutation was identified in a patient with exercise intolerance and developmental delay. A biochemical analysis revealed deficiencies in the activity of complex I (NADH:quinone oxidoreductase) and IV (cytochrome c oxidase) of this patient. Defects in complexes I and IV were confirmed in transmitochondrial cybrid cells containing the m.12955A > G mutation, suggesting that this mutation impairs complex I assembly, resulting in reduced stability of complex IV. Further functional investigations revealed that mitochondria with the m.12955A > G mutation exhibited lower OXPHOS coupling respiration and adenosine triphosphate (ATP) generation. In addition, the cytotoxic effects, determined as reactive oxygen species (ROS) and lactate levels in the present study, increased in the cells carrying a higher m.12955A > G mutant load. In conclusion, we identified m.12955A > G as a mitochondrial disease-related mutation. Therefore, screening of m.12955A > G is advised for the diagnosis of patients with mitochondrial disease.

The activation of protein homeostasis protective mechanisms perhaps is not responsible for lifespan extension caused by deficiencies of mitochondrial proteins in C. elegans.

During aging the ability of organisms to maintain the protein homeostasis declines and damaged and misfolded proteins accumulate in cells. But whether the deterioration of protein homeostasis is the cause or consequence of aging is not clearly understood. Mitochondrial dysfunctions usually lead to increased longevity in Caenorhabditis elegans, the cause of which is believed to be the activation of protein homeostasis protective mechanisms including mitochondrial unfolded protein response (UPR(mt)) and GCN-2 kinase mediated nutrient-sensing pathway. However, we investigated four genes which encode well-defined mitochondrial proteins and found that: (i) UPR(mt) activation was associated with not only increased longevity by knockdown of mfn-1, cco-1, or nuo-6, but also decreased longevity by mev-1 RNAi; (ii) The blockage of UPR(mt) pathway did not repress mfn-1, cco-1, or nuo-6 RNAi induced lifespan extension; (iii) The activation of UPR(mt) did not increase longevity; (iv) Knockdown of mfn-1, cco-1, or nuo-6 increased longevity independently of GCN-2. The combined results indicate that two important kinds of the protein homeostasis protective mechanisms, namely UPR(mt) and GCN-2 pathways, are not responsible for mitochondrial deficiency induced lifespan extension. The enhanced protection of protein homeostasis may be insufficient to slow aging, and there may be other mechanisms that contribute to the increased longevity in response to mitochondrial dysfunctions.