Drug-subphenotype interactions for cancer in type 2 diabetes mellitus.

On the basis of data obtained from a prospective cohort of Chinese patients with type 2 diabetes mellitus (T2DM), we discuss cancer subphenotypes (risk factors) in patients with T2DM, which can lead to drug-cancer subphenotype interactions. These subphenotypes include HDL cholesterol levels <1.0 mmol/l, co-occurrence of LDL cholesterol levels <2.8 mmol/l and triglyceride levels <1.7 mmol/l, and co-occurrence of LDL cholesterol levels <2.8 mmol/l and albuminuria. The increased risk of cancer associated with low levels of HDL cholesterol, low LDL cholesterol levels plus low triglyceride levels, and low levels of LDL cholesterol plus albuminuria can be reduced by treatment with metformin, renin-angiotensin system (RAS) inhibitors and statins, respectively. Mechanistic studies support the hypothesis that dysregulation of the 5'-AMP-activated protein kinase pathway and crosstalk between the RAS and insulin-like growth factor 1-cholesterol pathways create a cancer-promoting milieu in patients with T2DM. These findings highlight that in Chinese individuals, multiple pathways are implicated in the link between T2DM and cancer, which can generate multiple subphenotypes as well as drug-subphenotype interactions.

Differential inhibition of mitochondrial respiratory complexes by inhalation of combustion smoke and carbon monoxide, in vivo, in the rat brain.

Combustion smoke contains gases and particulates, which act via hypoxia and cytotoxicity producing mechanisms to injure cells and tissues. While carbon monoxide (CO) is the major toxicant in smoke, its toxicity is exacerbated in the presence of other compounds. Here, we examined modulations of mitochondrial and cytosolic energy metabolism by inhalation of combustion smoke versus CO, in vivo, in the rat brain. Measurements revealed reduced activities of respiratory chain (RC) complexes, with greater inhibition by smoke than equivalent CO in ambient air. In the case of RC complex IV, inhibition by CO and smoke was similar--suggesting that complex IV inhibition is primarily by the action of CO. In contrast, inhibition of complexes I and III was greater by smoke. Increases in cytosolic lactate dehydrogenase and pyruvate kinase activities accompanied inhibition of RC complexes, likely reflecting compensatory increases in cytosolic energy production. Together, the data provide new insights into the mechanisms of smoke inhalation-induced perturbations of brain energetics, which impact neuronal function and contribute to the development of neuropathologies in survivors of exposures to CO and combustion smoke.

Impaired mitochondrial respiration and protein nitration in the rat hippocampus after acute inhalation of combustion smoke.

Survivors of massive inhalation of combustion smoke endure critical injuries, including lasting neurological complications. We have previously reported that acute inhalation of combustion smoke disrupts the nitric oxide homeostasis in the rat brain. In this study, we extend our findings and report that a 30-minute exposure of awake rats to ambient wood combustion smoke induces protein nitration in the rat hippocampus and that mitochondrial proteins are a sensitive nitration target in this setting. Mitochondria are central to energy metabolism and cellular signaling and are critical to proper cell function. Here, analyses of the mitochondrial proteome showed elevated protein nitration in the course of a 24-hour recovery following exposure to smoke. Mass spectrometry identification of several significantly nitrated mitochondrial proteins revealed diverse functions and involvement in central aspects of mitochondrial physiology. The nitrated proteins include the ubiquitous mitochondrial creatine kinase, F1-ATP synthase alpha subunit, dihydrolipoamide dehydrogenase (E3), succinate dehydrogenase Fp subunit, and voltage-dependent anion channel (VDAC1) protein. Furthermore, acute exposure to combustion smoke significantly compromised the respiratory capacity of hippocampal mitochondria. Importantly, elevated protein nitration and reduced mitochondrial respiration in the hippocampus persisted beyond the time required for restoration of normal oxygen and carboxyhemoglobin blood levels after the cessation of exposure to smoke. Thus, the time frame for intensification of the various smoke-induced effects differs between blood and brain tissues. Taken together, our findings suggest that nitration of essential mitochondrial proteins may contribute to the reduction in mitochondrial respiratory capacity and underlie, in part, the brain pathophysiology after acute inhalation of combustion smoke.

A possible role for hypoxia-induced apelin expression in enteric cell proliferation.

Apelin is the endogenous ligand for the APJ receptor, and apelin and APJ are expressed in the gastrointestinal (GI) tract. Intestinal inflammation increases intestinal hypoxia-inducible factor (HIF) and apelin expression. Hypoxia and inflammation are closely linked cellular insults. The purpose of these studies was to investigate the influence of hypoxia on enteric apelin expression. Exposure of rat pups to acute hypoxia increased hepatic, stomach-duodenal, and colonic apelin mRNA levels 10-, 2-, and 2-fold, respectively (P < 0.05 vs. controls). Hypoxia also increased colonic APJ mRNA levels, and apelin treatment during hypoxia exposure enhanced colonic APJ mRNA levels further. In vitro hypoxia also increased apelin and APJ mRNA levels. The hypoxia-induced elevation in apelin expression is most likely mediated by HIF, since HIF-activated apelin transcriptional activity is dependent on an intact, putative HIF binding site in the rat apelin promoter. Acute exposure of rat pups to hypoxia lowered gastric and colonic epithelial cell proliferation; hypoxia in combination with apelin treatment increased epithelial proliferation by 50%. In vitro apelin treatment of enteric cells exposed to hypoxia increased cell proliferation. Apelin treatment during normoxia was ineffective. Our studies imply that the elevation in apelin expression during hypoxia and inflammation in the GI tract functions in part to stimulate epithelial cell proliferation.

Sustained hypoxia modulates mitochondrial DNA content in the neonatal rat brain.

The effects of placental insufficiency and preterm birth on neurodevelopment can be modeled in experimental settings of neonatal hypoxia in rodents. Here, rat pups were reared in reduced oxygen (9.5%) for 11 days, starting on postnatal day 3 (P3). This led to a significant reduction in brain and body weight gain in hypoxic pups compared to age-matched normoxia-reared controls, plausibly reflecting an inability to fulfill the energetic needs of normal growth and development. Adaptive processes designed to augment energetic capacity in eukaryotes include stimulation of mitochondrial biogenesis. We show that after 11 days of sustained hypoxia, the levels of nuclear respiratory factor-1 and mitochondrial transcription factor A are elevated and the content of mitochondrial DNA (mtDNA) is greater in the hypoxic P14 pup brain compared to normoxic conditions. Corresponding immunohistochemical analyses reveal increased density of mtDNA in large cortical neurons. In contrast, no changes in mtDNA content are observed in the brain of pups reared for 24 h (P3-P4) under hypoxic conditions. Together, these data suggest that prolonged inadequate oxygenation may trigger a compensatory increase in neuronal mitochondrial DNA content to partially mitigate compromised energy homeostasis and reduced energetic capacity in the developing hypoxic brain.

Accumulation of oxidatively generated DNA damage in the brain: a mechanism of neurotoxicity.

Unrepaired or erroneously repaired DNA lesions drive genomic instability and contribute to cellular and organ decline. Since delayed neuropathologies are common in survivors of smoke inhalation injuries, we asked whether the integrity of brain DNA might be compromised by acute exposure to combustion smoke. Although many studies demonstrate that the brain is equipped to repair oxidatively damaged DNA, to date, the capacity for accurate DNA repair under conditions of disrupted oxygenation and oxidative stress has not been defined. We show that DNA adducts detectable by their ability to block PCR amplification form in the rat hippocampus after acute exposure to smoke. To identify the different types of adducts and to dissect their temporal formation and repair profiles in vivo in the brain, we used DNA-modifying enzymes to convert specific adducts into strand breaks prior to PCR amplification. Using this strategy, we detected formation of oxidative DNA adducts early on after smoke inhalation, while mismatched bases emerged at the later recovery times, potentially due to an erroneous DNA repair process. Erroneous repair can be mutagenic and because the initial smoke-induced oxidative damage to DNA is extensive, compromised fidelity of DNA repair may underlie neurotoxicity and contribute to delayed death of hippocampal neurons.

A rat model of smoke inhalation injury: influence of combustion smoke on gene expression in the brain.

Acute smoke inhalation causes death and injury in victims of home and industrial fires as well as victims of combat situations. The lethal factors in combustion smoke inhalation are toxic gases and oxygen deficiency, with carbon monoxide (CO) as a primary cause of death. In survivors, inhalation of smoke can result in severe immediate and delayed neuropathologies. To gain insight into the progression of molecular events contributing to smoke inhalation sequelae in the brain, we developed a smoke inhalation rat model and conducted a genome-wide analysis of gene expression. Microarray analysis revealed a modified brain transcriptome with changes peaking at 24 h and subsiding within 7 days post-smoke. Overall, smoke inhalation down regulated genes associated with synaptic function, neurotransmission, and neurotrophic support, and upregulated genes associated with stress responses, including nitric oxide synthesis, antioxidant defenses, proteolysis, inflammatory response, and glial activation. Notably, among the affected genes, many have been previously implicated in other types of brain injury, demonstrating the usefulness of microarrays for analysis of changes in gene expression in complex insults. In accord with previously described modulations of nitric oxide homeostasis in CO poisoning, microarray analysis revealed increased brain expression of nitric oxide synthase (NOS) and NOS ligand after inhalation of smoke. Furthermore, immunostaining showed significant elevations in perivascular NOS and in protein nitration, corroborating the involvement of nitric oxide perturbations in post-smoke sequelae in the brain. Thus, the new rat model, in combination with microarray analyses, affords insight into the complex molecular pathophysiology of smoke inhalation in the brain.

N-terminus of the rat adenine glycosylase MYH affects excision rates and processing of MYH-generated abasic sites.

Repair of most modified and mispaired bases in the genome is initiated by DNA glycosylases, which bind to their respective targets and cleave the N-glycosyl bond to initiate base excision repair (BER). The mammalian homolog of the Escherichia coli MutY DNA glycosylase (MYH) cleaves adenine residues paired with either oxidized or non-modified guanines. MYH is crucial for the avoidance of mutations resulting from oxidative DNA damage. Multiple N-terminal splice variants of MYH exist in mammalian cells and it is likely that different variants result in the production of enzymes with altered properties. To investigate whether modifications in the N-terminus are consequential to MYH function, we overexpressed intact and N-terminal-deletion rat MYH proteins and examined their activities. We found that deletion of 75 amino acids, which perturbs the catalytic core that is conserved with E.coli MutY, abolished excision activity. In contrast, deletions limited to the extended mammalian N-terminal domain, differentially influenced steady-state excision rates. Notably, deletion of 50 amino acids resulted in an enzyme with a significantly lower K(m) favoring formation of excision products with 3'-OH termini. Our findings suggest that MYH isoforms divergent in the N-terminus influence excision rates and processing of abasic sites.