TRDMT1 exhibited protective effects against LPS-induced inflammation in rats through TLR4-NF-κB/MAPK-TNF-αpathway

Background:Inflammation is a complex physiological and pathological process.Although many types of inflammation are well characterized,their physiological func-tions are largely unknown.tRNA aspartic acid methyltransferase 1(TRDMT1)has been implicated as a stress-related protein,but its intrinsic biological role is unclear.Methods:We constructed a Trdmt1 knockout rat and adopted the LPS-induced sepsis model.Survival curve,histopathological examination,expression of inflammatory fac-tors,and protein level of TLR4 pathway were analyzed.Results:Trdmt1 deletion had no obvious impact on development and growth.Trdmt1 de-letion slightly increased the mortality during aging.Our data showed that Trdmt1 strongly responded in LPS-treated rats,and Trdmt1 knockout rats were vulnerable to LPS treat-ment with declined survival rate.We also observed more aggravated tissue damage and more cumulative functional cell degeneration in LPS-treated knockout rats compared with control rats.Further studies showed upregulated TNF-αlevel in liver,spleen,lung,and serum tissues,which may be explained by enhanced p65 and p38 phosphorylation.Conclusions:Our data demonstrated that Trdmt1 plays a protective role in inflamma-tion by regulating the TLR4-NF-κB/MAPK-TNF-αpathway.This work provides useful information to understand the TRDMT1 function in inflammation.

SARS-CoV-2 infection aggravates chronic comorbidities of cardiovascular diseases and diabetes in mice

Background:Cardiovascular diseases(CVDs)and diabetes mellitus(DM)are top two chronic comorbidities that increase the severity and mortality of COVID-19.However,how SARS-CoV-2 alters the progression of chronic diseases remain unclear.Methods:We used adenovirus to deliver h-ACE2 to lung to enable SARS-CoV-2 infection in mice.SARS-CoV-2’s impacts on pathogenesis of chronic diseases were studied through histopathological,virologic and molecular biology analysis.Results:Pre-existing CVDs resulted in viral invasion,ROS elevation and activation of apoptosis pathways contribute myocardial injury during SARS-CoV-2 infection.Viral infection increased fasting blood glucose and reduced insulin response in DM model.Bone mineral density decreased shortly after infection,which associated with impaired PI3K/AKT/mTOR signaling.Conclusion:We established mouse models mimicked the complex pathological symptoms of COVID-19 patients with chronic diseases.Pre-existing diseases could impair the inflammatory responses to SARS-CoV-2 infection,which further aggravated the pre-existing diseases.This work provided valuable information to better understand the interplay between the primary diseases and SARS-CoV-2 infection.

Knock in of a hexanucleotide repeat expansion in the C9orf72 gene induces ALS in rats

Background:The GGGGCC(G4C2)repeat expansion in the human open reading frame 72 on chromosome 9,C9orf72,is the most common cause of amyotrophic lateral sclerosis(ALS).Studies in transgenic mouse models have linked the pathogenic mechanism of G4C2 repeat expansion to RNA foci or the accumulation of unnatural dipeptide repeats in neurons.However,only one of the existing transgenic mouse lines developed typical ALS.Methods:C9orf72 knockin rats were generated by knockin of 80 G4C2 repeats with human flanking fragments within exon1a and exon1b at the rat C9orf72 locus.Protein expression was detected by western blot.Motor coordination and grip force were measured using a Rotarod test and a grip strength test.Neurodegeneration was assessed by Nissl staining with cresyl violet.Results:C9orf72 haploinsufficiency reduced C9orf72 protein expression 40%in the cerebrum,cerebellum and spinal cords from knockin rats(P<.05).The knockin(KI)rats developed motor deficits from 4 months of age.Their falling latencies and grip force were decreased by 67%(P<.01)and 44%(P<.01),respectively,at 12 months of age compared to wild-type(WT)mice.The knockin of the hexanucleotide repeat expansion(HRE)caused a 47%loss of motor neurons in the spinal cord(P<.001)and 25%(5/20)of female KI rats developed hind limb paralysis at 13 to 24 months.Conclusion:Motor defects in KI rats may result from neurotoxicity caused by HRE and the resulting reduction in C9orf72 protein due to haploinsufficiency.These KI rats could be a useful model for investigating the contributions of loss-of-function to neurotoxicity in C9orf72-related ALS.

Novel rat model of multiple mitochondrial dysfunction syndromes(MMDS)complicated with cardiomyopathy

Background:Multiple mitochondrial dysfunction syndromes(MMDS)presents as complex mitochondrial damage,thus impairing a variety of metabolic pathways.Heart dysplasia has been reported in MMDS patients;however,the specific clinical symptoms and pathogenesis remain unclear.More urgently,there is a lack of an animal model to aid research.Therefore,we selected a reported MMDS causal gene,Isca1,and established an animal model of MMDS complicated with cardiac dysplasia.Methods:The myocardium-specific Isca1 knockout heterozygote(Isca1 HET)rat was obtained by crossing the Isca1 conditional knockout(Isca1 cKO)rat with theαmyosin heavy chain Cre(α-MHC-Cre)rat.Cardiac development characteristics were determined by ECG,blood pressure measurement,echocardiography and histopatho-logical analysis.The responsiveness to pathological stimuli were observed through adriamycin treatment.Mitochondria and metabolism disorder were determined by activity analysis of mitochondrial respiratory chain complex and ATP production in myocardium.Results:ISCA1 expression in myocardium exhibited a semizygous effect.Isca1 HET rats exhibited dilated cardiomyopathy characteristics,including thin-walled ventri-cles,larger chambers,cardiac dysfunction and myocardium fibrosis.Downregulated ISCA1 led to deteriorating cardiac pathological processes at the global and organiza-tional levels.Meanwhile,HET rats exhibited typical MMDS characteristics,including damaged mitochondrial morphology and enzyme activity for mitochondrial respira-tory chain complexesⅠ,ⅡandⅣ,and impaired ATP production.Conclusion:We have established a rat model of MMDS complicated with cardiomyopathy,it can also be used as model of myocardial energy metabolism dysfunction and mitochondrial cardiomyopathy.This model can be applied to the study of the mechanism of energy metabolism in cardiovascular diseases,as well as research and development of drugs.

Knockout of ISCA 1 causes early embryonic death in rats

Background : Iron-sulfur cluster assembly 1( ISCA 1) is an iron-sulfur(Fe/S) carrier protein that accepts Fe/S from a scaffold protein and transfers it to target proteins including the mitochondrial Fe/S containing proteins. ISCA 1 is also the newly identified causal gene for multiple mitochondrial dysfunctions syndrome( MMDS). However, our knowledge about the physiological function of ISCA 1 in vivo is currently limited. In this study, we generated an ISCA 1 knockout rat line and analyzed the embryo development. Methods : ISCA 1 knockout rats were generated by replacing the exon1 of ISCA 1 gene with the mC herry-Cre fusion gene using CRISPR-Cas9 technology. The ISCA 1 expression pattern was analyzed by fluorescence imaging using ISCA 1 promotor driven Cre and mC herry expression. The embryonic morphology was examinated by microscope and mitochondrial proteins were tested by Western blot. Results : An ISCA 1 knockout rat line was obtained, which expressed mC herry-Cre fusion protein. Both of the fluorescence images from mC herry and Cre induced mC herry in a reporter rat strain, showing that ISCA 1 expressed in most of the tissues in rats. The ISCA 1 knockout resulted in abnormal development at 8.5 days, with a significant decrease of NDUFA 9 protein and an increase of aconitase 2( ACO 2) in rat embryos. Conclusion : Deletion of ISCA 1 induced early death in rats. ISCA 1 affected the expression of key proteins in the mitochondrial respiratory chain complex, suggesting that ISCA 1 has an important influence on the respiratory complex and energy metabolism.

The first genetically gene-edited babies: It's “irresponsible and too early”

A scientist, Jiankui He of Southern University of Science and Technology of China,recently claimed at the Second International Summit on Human Genome Editing in Hong Kong on 29 November that he has created the world's first genetically altered babies using CRISPR. This announcement sparked controversy and criticism. The newly developed CRISPR/Cas9 technique has been applied to genetic modification of many kinds of animals. However, the technique is still in its infancy and many questions remain to be answered before it can be used for clinical purposes, especially for reproductive purposes.