Suppressed mitochondrial respiration via NOX5-mediated redox imbalance contributes to the antitumor activity of anlotinib in oral squamous cell carcinoma

Anlotinib,a novel multitarget tyrosine kinase inhibitor,has shown promising results in the management of various carcinomas.This study aimed to investigate the antitumor activity of anlotinib in oral squamous cell carcinoma(OSCC)and the underlying molecular mechanism.A retrospective clinical study revealed that anlotinib improved the median progression-free survival(m PFS)and median overall survival(m OS)of patients with recurrent and metastatic(R/M)OSCC,respectively.Functional studies revealed that anlotinib markedly inhibited in vitro proliferation of OSCC cells and impeded in vivo tumor growth of OSCC patientderived xenograft models.Mechanistically,RNA-sequencing identified that oxidative stress,oxidative phosphorylation and AKT/m TOR signaling were involved in anlotinib-treated OSCC cells.Anlotinib upregulated NADPH oxidase 5(NOX5)expression,elevated reactive oxygen species(ROS)production,impaired mitochondrial respiration,and promoted apoptosis.Moreover,anlotinb also inhibited phosphoAkt(p-AKT)expression and elevated p-e IF2αexpression in OSCC cells.NOX5 knockdown attenuated these inhibitory effects and cytotoxicity in anlotinib-treated OSCC cells.Collectively,we demonstrated that anlotinib monotherapy demonstrated favorable anticancer activity and manageable toxicities in patients with R/M OSCC.The antitumor activity of anlotinib in OSCC may be mainly involved in the suppression of mitochondrial respiration via NOX5-mediated redox imbalance and the AKT/e IF2αpathway.

The profiles of mitochondrial respiration and glycolysis using extracellular flux analysis in porcine enterocyte IPEC-J2

The porcine intestinal mucosa require large amounts of energy for nutrient processing and cellular functions and is vulnerable to injury by weaning stress involving bioenergetics failure. The mitochondrial bioenergetic measurement in porcine enterocytes have not been defined. The present study was to establish a method to measure mitochondrial respiratory function and profile mitochondrial function of IPEC-J2 using cell mito stress test and glycolysis stress test assay by XF24 extracellular flux analyzer. The optimal seeding density and concentrations of the injection compounds were determined to be 40,000 cells/well as well as 0.5 μ M oligomycin, 1 μM carbonyl cyanide p-trifluoromethoxy-phenylhydrazone(FCCP) and 1 μM rotenone & antimycin A, respectively. The profiles of mitochondrial respiration and glycolysis confirmed that porcine enterocyte preferentially derived much more energy from glutamine than glucose. These results will provide a basis for further study of mitochondrial function and bioenergetics of the porcine small intestine.

Targeting mitochondrial transcription factor A sensitizes pancreatic cancer cell to gemcitabine

Background:The survival of pancreatic cancer cells,particularly cancer stem cells which are responsible for tumor relapse,depends on mitochondrial function.Mitochondrial transcription factor A(TFAM)is critical for the regulation of mitochondrial DNA and thus mitochondrial function.However,the possible involvement of TFAM in pancreatic cancer is unknown.Methods:Human samples were obtained from pancreatic cancers and their adjacent tissues;human pancreatic cell lines were cultured in RPMI1640 medium.TFAM expressions in pancreatic tissues and cultured cells were determined using immunohistochemistry,ELISA,and reverse transcription polymerase chain reaction(RT-PCR).The effect of TFAM on cell growth,migration,colony formation and apoptosis were evaluated.Mitochondrial biogenesis in pancreatic cancer and normal cells were examined.Results:The majority of pancreatic cancer tissues exhibited higher TFAM expression compared to the adjacent counterparts.Consistently,TFAM mRNA and protein levels were higher in pancreatic cancer cell lines than in immortalized normal pancreatic epithelial cells.There was no difference on TFAM level between gemcitabine-sensitive and resistant pancreatic cancer cells.Functional analysis demonstrated that TFAM overexpression activated pancreatic normal and tumor cells whereas TFAM inhibition effectively inhibited the growth of pancreatic cancer cells.TFAM inhibition enhanced gemcitabine’s cytotoxicity and suppressed growth,anchorage-independent colony formation and survival of gemcitabine-resistant pancreatic cancer cells.Mechanistic studies showed that TFAM inhibition resulted in remarkable mitochondrial dysfunction and energy crisis followed by oxidative stress.The basal mitochondrial biogenesis level correlated well with TFAM level in pancreatic cancer cells.Conclusions:TFAM played essential roles in pancreatic cancer via regulating mitochondrial functions which highlighted the therapeutic value of inhibiting TFAM to overcome gemcitabine resistance.

Steatotic livers are susceptible to normothermic ischemiareperfusion injury from mitochondrial Complex-Ⅰ?dysfunction

AIM: To assess the effects of ischemic preconditioning(IPC, 10-min ischemia/10-min reperfusion) on steatotic liver mitochondrial function after normothermic ischemia-reperfusion injury(IRI).METHODS: Sixty male Sprague-Dawley rats were fed8-wk with either control chow or high-fat/high-sucrose diet inducing > 60% mixed steatosis. Three groups(n = 10/group) for each dietary state were tested:(1) the IRI group underwent 60 min partial hepatic ischemia and 4 h reperfusion;(2) the IPC group underwent IPC prior to same standard IRI; and(3) sham underwent t h e s a m e s u r g e r y w i t h o u t I R I o r I P C. H e p a t i c mitochondrial function was analyzed by oxygraphs. Mitochondrial Complex-Ⅰ, Complex-Ⅱ enzyme activity, serum alanine aminotransferase(ALT), and histological injury were measured.RESULTS: Steatotic-IRI livers had a greater increase in ALT(2476 ± 166 vs 1457 ± 103 IU/L, P < 0.01) and histological injury following IRI compared to the lean liver group. Steatotic-IRI demonstrated lower Complex-Ⅰ?activity at baseline [78.4 ± 2.5 vs 116.4 ± 6.0 nmol/(min.mg protein), P < 0.001] and following IRI [28.0 ± 6.2 vs 104.3 ± 12.6 nmol/(min.mg protein), P < 0.001]. Steatotic-IRI also demonstrated impaired Complex-Ⅰ?function post-IRI compared to the lean liver IRI group. Complex-Ⅱ activity was unaffected by hepatic steatosis or IRI. Lean liver mitochondrial function was unchanged following IRI. IPC normalized ALT and histological injury in steatotic livers but had no effect on overall steatotic liver mitochondrial function or individual mitochondrial complex enzyme activities. CONCLUSION: Warm IRI impairs steatotic liver Complex-Ⅰ?activity and function. The protective effects of IPC in steatotic livers may not be mediated through mitochondria.

Commentary: Inhibitors of mitochondrial respiratory chain in the treatment of type 2 diabetes

Prevalence of type 2 diabetes has been increased worldwide following the high incidence of obesity and growing population of aging, which are two conditions for absolute energy excess with accumulation of TAG in the body and relative energy excess due to reduced energy expense, respectively. It appears that type 2 diabetes is a compensatory mechanism of body to energy excess through urine discharge of glucose.

Liver mitochondrial and whole-animal level metabolic compensation in a catfish during seasonal acclimatization

To examine whether metabolic compensation during seasonal acclimatization at the liver mitochondrial level is consistent with that at the whole-animal level, respiration rates of liver mitochondria and resting metabolic rates in winter- and sum- meracclimatized southern catfish （Silurus meridionalis Chen） were measured. At 12.5, 17.5, 22.5, 27.5 and 32.5~C, the mean values of state 3 respiration rates were 12.21, 13.84, 18.96, 24.78 and 32.01 nmol O2min-1 mg-1 mitochondrial protein in the winter group, and 8.56, 9.20, 17.32, 22.74 and 26.32 nmol 02 min-1 mgq in the summer group, respectively. At the five assay temperatures the resting metabolic rates were 24.86, 42.68, 61.59, 84.10 and 125.65 mg 02 h-1 kgI body mass in the winter group, and 22.89, 40.59, 52.94, 75.13 and 109.35 mg Oz h-1 kg-1 in the summer group, respectively. Total mitochondrial respiration rates in the liver organ were estimated based on state 3 respiration rates, mitochondrial protein content and organ mass, and the mean values were 72.96, 71.87, 112.47, 167.35 and 183.27 nmol Ozmin-lin the winter group, and were 47.89, 47.39, 105.67, 138.18 and 132.29 nmol 02 min-1 in the summer group, respectively. Metabolic compensation caused by seasonal acclimatization occurred at the liver mitochondrial level and compensation at the liver organ level was found to be more efficient because of an in- crease in metabolic capacity of mitochondria and a boost in organ mass. Metabolic compensation at the whole-animal level was not detected. During seasonal acclimatization, the effect of metabolic compensation at liver mitochondrial level is inconsistent with that at the whole-animal level in the southern catfish. This may be due to different degrees of regulation of metabolic mechanisms among various tissues and organs in an acclimatized organism