Recurrent glucose deprivation leads to the preferential use of lactate by neurons in the ventromedial hypothalamus.

Increased GABAergic output in the ventromedial hypothalamus (VMH) contributes to counterregulatory failure in recurrently hypoglycemic (RH) rats, and lactate, an alternate fuel source in the brain, contributes to this phenomenon. The current study assessed whether recurring bouts of glucose deprivation enhanced neuronal lactate uptake and, if so, whether this influenced γ-aminobutyric acid (GABA) output and the counterregulatory responses. Glucose deprivation was induced using 5-thioglucose (5TG). Control rats received an infusion of artificial extracellular fluid. These groups were compared with RH animals. Subsequently, the rats underwent a hypoglycemic clamp with microdialysis. To test whether 5TG affected neuronal lactate utilization, a subgroup of 5TG-treated rats was microinjected with a lactate transporter inhibitor [cyano-4-hydroxycinnamate (4CIN)] just before the start of the clamp. Both RH and 5TG raised VMH GABA levels, and this was associated with impaired counterregulatory responses. 4CIN reduced VMH GABA levels and restored the hormone responses in the 5TG group. We then evaluated [14C]lactate uptake in hypothalamic neuronal cultures. Recurring exposure to low glucose increased monocarboxylate transporter-2 mRNA expression and augmented lactate uptake. Taken together, our data suggest that glucose deprivation, per se, enhances lactate utilization in hypothalamic neurons, and this may contribute to suppression of the counterregulatory responses to hypoglycemia.

Propionate and butyrate induce gene expression of monocarboxylate transporter 4 and cluster of differentiation 147 in cultured rumen epithelial cells derived from preweaning dairy calves.

Short-chain fatty acids (SCFAs) are the main source of energy for postweaning ruminants. The monocarboxylic acid transporters, MCT1 and MCT4, are thought to contribute to the absorption of SCFAs from the surface of the rumen following weaning. The present study measured changes in MCT1 and MCT4 expression in ruminal epithelial cells isolated from male preweaning (22 to 34 d old, n = 6) and postweaning (55 to 58 d old, n = 8) calves after euthanasia and sought to examine whether SCFAs stimulate the expression of these transporters. In the current study, cluster of differentiation 147 (CD147) gene expression in the rumen was also investigated since CD147 has been considered to act as ancillary protein for MCT1 and MCT4 to express their correct function. The gene expression levels of MCT1, MCT4, and CD147 in the rumen were found to be significantly higher in postweaning calves than in preweaning calves. Strong MCT1 immunoreactivity was detected in both the stratum basale (SB) and the stratum spinosum (SS) in postweaning ruminal epithelium. Expression of MCT1 in preweaning calves was localized to a specific region of the SB and of the SS. MCT4-immunopositive cells were detected in the stratum corneum (SC) of the ruminal epithelium in postweaning calves. However, only a low level of signal was detected in the SC of preweaning animals. Furthermore, in vitro experiments, ruminal epithelial cells were incubated for 24 h with acetate (0.04, 0.4, and 4 mM), propionate (0.2, 2, and 20 mM), butyrate (0.1, 1, and 10 mM), or ß-hydroxybutyrate (BHBA; 0.1, 1, and 10 mM), respectively. Both propionate and butyrate induced an increase in the gene expression levels of MCT4 and CD147, but did not affect MCT1 gene expression. There are no significant effects of acetate and BHBA treatment on these gene expressions. Taken together, these results suggest that an increase in MCT4 and CD147 gene expression in the ruminal epithelium of postweaning calves is likely to be due to the effects of propionate and butyrate derived from a solid-based diet, which may contribute to ruminal development following weaning.

Sorafenib-Induced Changes in Thyroid Hormone Levels in Patients Treated for Hepatocellular Carcinoma.

Context: The pathogenesis of tyrosine kinase inhibitor-induced thyroid hormone (TH) alterations are still a matter of debate. Objective: The objective of this study was to determine the effects of sorafenib on TH levels in patients with hepatocellular carcinoma (HCC) and to evaluate possible mechanisms. Design: We performed a prospective cohort study between 2009 and 2016. Setting: This study was conducted at a tertiary referral center. Patients: This study included 57 consecutive patients with HCC who were treated with sorafenib. Main Outcome Measure: Thyroid-stimulating hormone (TSH) and free thyroxine (FT4) levels were measured every 6 weeks, and extensive thyroid function tests (TFTs) were measured before treatment (t0), after 6 weeks (t6), and at the end of therapy. The effect of sorafenib on TH transport by monocarboxylate transporter (MCT)8 or MCT10 was tested in transfected COS1 cells. Results: Four patients (7%) developed thyroiditis. Among the other patients, 30% had elevation of TSH or FT4 above the normal range. Overall, between t0 and t6, mean TSH increased from 1.28 to 1.57 mU/L (P < 0.001) and mean FT4 from 18.4 to 21.2 pmol/L (P < 0.001). Simultaneously, the serum triiodothyronine (T3)/reverse triiodothyronine ratio and the (T3/thyroxine) ×100 ratio decreased. Sorafenib decreased cellular T3 uptake by MCT8 and to a lesser extent by MCT10. Conclusions: These in vivo data suggest that sorafenib affects TFTs on multiple levels. Our in vitro experiments suggest a possible role of sorafenib-induced inhibition of T3 transport into the cell by MCT8 and MCT10.

Association Between SLC16A5 Genetic Variation and Cisplatin-Induced Ototoxic Effects in Adult Patients With Testicular Cancer.

IMPORTANCE: Cisplatin-induced ototoxic effects are an important complication that affects testicular cancer survivors as a consequence of treatment. The identification of genetic variants associated with this adverse drug reaction will further our mechanistic understanding of its development and potentially lead to strategies to prevent ototoxic effects. OBJECTIVE: To identify the genetic variants associated with cisplatin-induced ototoxic effects in adult testicular cancer patients. DESIGN, SETTING, AND PARTICIPANTS: This retrospective study was performed by the Canadian Pharmacogenomics Network for Drug Safety using patients recruited from 5 adult oncology treatment centers across Canada. Male patients who were 17 years or older, diagnosed with germ cell testicular cancer, and previously treated with cisplatin-based chemotherapy were recruited from July 2009 to April 2013 using active surveillance methodology. Cisplatin-induced ototoxic effects were independently diagnosed by 2 audiologists. Patients were genotyped for 7907 variants using a custom pharmacogenomic array. Logistic regression was used to identify genetic variants that were significantly associated with ototoxic effects. The validity of these findings was confirmed through independent replication and cell-based functional assays. EXPOSURES: Cisplatin-based chemotherapy. MAIN OUTCOMES AND MEASURES: Cisplatin-induced ototoxic effects. RESULTS: After exclusions, 188 patients (median [interquartile range] age, 31 [24-39] years) were enrolled in this study to form the discovery and replication cohorts. Association and fine-mapping analyses identified a protein-coding variant, rs4788863 in SLC16A5, that was associated with protection against cisplatin-induced ototoxic effects in 2 independent cohorts (combined cohort: odds ratio, 0.06; 95% CI, 0.02-0.22; P = 2.17 × 10-7). Functional validation of this transporter gene revealed that in vitro SLC16A5-silencing altered cellular responses to cisplatin treatment, supporting a role for SLC16A5 in the development of cisplatin-induced ototoxic effects. These results were further supported by the literature, which provided confirmatory evidence for the role that SLC16A5 plays in hearing. CONCLUSIONS AND RELEVANCE: This study has identified a novel association between protein-coding variation in SLC16A5 and cisplatin-induced ototoxic effects. These findings have provided insight into the molecular mechanisms of this adverse drug reaction in adult patients with germ cell testicular cancer. Given that previous studies have shown that cimetidine, an SLC16A5-inhibitor, prevents murine cisplatin-induced ototoxic effects, the findings from this study have important implications for otoprotectant strategies in humans.

Efficient Activation of Pathogenic ΔPhe501 Mutation in Monocarboxylate Transporter 8 by Chemical and Pharmacological Chaperones.

Monocarboxylate transporter 8 (MCT8) is a thyroid hormone transmembrane transporter expressed in many cell types, including neurons. Mutations that inactivate transport activity of MCT8 cause severe X-linked psychomotor retardation in male patients, a syndrome originally described as the Allan-Herndon-Dudley syndrome. Treatment options currently explored the focus on finding thyroid hormone-like compounds that bypass MCT8 and enter cells through different transporters. Because MCT8 is a multipass transmembrane protein, some pathogenic mutations affect membrane trafficking while potentially retaining some transporter activity. We explore here the effects of chemical and pharmacological chaperones on the expression and transport activity of the MCT8 mutant ΔPhe501. Dimethylsulfoxide, 4-phenylbutyric acid as well as its sodium salt, and the isoflavone genistein increase T3 uptake into MDCK1 cells stably transfected with mutant MCT8-ΔPhe501. We show that ΔPhe501 represents a temperature-sensitive mutant protein that is stabilized by the proteasome inhibitor MG132. 4-Phenylbutyrate has been used to stabilize ΔPhe508 mutant cystic fibrosis transmembrane conductance regulator protein and is in clinical use in patients with urea cycle defects. Genistein is enriched in soy and available as a nutritional supplement. It is effective in stabilizing MCT8-ΔPhe501 at 100 nM concentration. Expression of the L471P mutant is increased in response to phenylbutyrate, but T3 uptake activity is not induced, supporting the notion that the chaperone specifically increases membrane expression. Our findings suggest that certain pathogenic MCT8 mutants may be responsive to (co-)treatment with readily available compounds, which increase endogenous protein function.

Functional cooperation of SMCTs and URAT1 for renal reabsorption transport of urate.

  Urate is mainly excreted into urine in humans. Serum urate level is regulated by a urate transport system located on the renal proximal tubule. Urate transporter 1 (URAT1) is located on the apical side of the renal proximal tubule and is responsible for the reabsorption of urate from the luminal side into tubular cells. At the same site, it has been hypothesized that sodium-coupled monocarboxylate transporters (SMCTs) are responsible for the transportation of monocarboxylates such as lactate and nicotinate, which are exchanged for urate transport via URAT1. Accordingly, SMCTs could enhance URAT1-mediated urate reabsorption by providing monocarboxylates for the exchange. The present study was carried out to clarify the hypothesized functional cooperative relationship between URAT1 and SMCTs in the reabsorptive transport of urate. By preloading nicotinate in SMCT1/URAT1-coexpressing Xenopus oocytes, URAT1-mediated urate transport was stimulated. Nicotinate was taken up by SMCT1 but not by URAT1. When removing sodium ions from the uptake medium, the stimulation effect was decreased. When adding SMCT1 inhibitors, the stimulation effect was also reduced. The results from this study indicate the cooperative relationship of URAT1 and SMCT1, and that SMCT1 is a potential target for the alteration of renal handling of urate indirectly.

Increased densities of monocarboxylate transport protein MCT1 after chronic administration of nicotine in rat brain.

Chronic administration of nicotine is followed by a general stimulation of brain metabolism that results in a distinct increase of glucose transport protein densities for Glut1 and Glu3, and local cerebral glucose utilization (LCGU). This increase of LCGU might be paralleled by an enhanced production of lactate. Therefore, the question arose as to whether chronic nicotine infusion is accompanied by increased local densities of monocarboxylate transporter MCT1 in the brain. Secondly, we inquired whether LCGU might be correlated with local densities of MCT1 during normal conditions and after chronic nicotine infusion. Nicotine was given subcutaneously for 1 week by osmotic mini-pumps and local densities of MCT1 were measured by immunoautoradiographic methods in cryosections of rat brains. MCT1 density was significantly increased in 21 of 32 brain structures investigated (median increase 15.0+/-3.6%). Immunohistochemical stainings of these substructures revealed an over-expression of MCT1 within endothelial cells and astrocytes of treated animals. A comparison of 23 MCT1 densities with LCGU measured in the same structures in a previous study revealed a partial correlation between both parameters under control conditions and after chronic nicotine infusion. 10 out of 23 brain areas, which showed a significant increase of MCT1 density due to chronic nicotine infusion, also showed a significant increase of LCGU. In summary, our data show that chronic nicotine infusion induces a moderate increase of local and global density of MCT1 in defined brain structures. However, in terms of brain topologies and substructures this phenomenon did partially match with increased LCGU. It is concluded that MCT1 transporters were upregulated during chronic nicotine infusion at the level of brain substructures and, at least partially, independently of LCGU.

Nutrient transporters in cancer: relevance to Warburg hypothesis and beyond.

Tumor cells have an increased demand for nutrients; this demand is met by increased availability of nutrients through vasculogenesis and by enhanced cellular entry of nutrients through upregulation of specific transporters. This review focuses on three groups of nutrient transporters relevant to cancer: glucose transporters, lactate transporters, and amino acid transporters. Tumor cells enhance glucose uptake via induction of GLUT1 and SGLT1, and coordinate the increased entry of glucose with increased glycolysis. Since enhanced glycolysis in cancer is associated with lactate production, tumor cells must find a way to eliminate lactic acid to prevent cellular acidification. This is achieved by the upregulation of MCT4, a H+-coupled lactate transporter. In addition, the Na+-coupled lactate transporter SMCT1 is silenced in cancer. SMCT1 also transports butyrate and pyruvate, which are inhibitors of histone deacetylases. The silencing of SMCT1 occurs in cancers of a variety of tissues. Re-expression of SMCT1 in cancer cell lines leads to growth arrest and apoptosis in the presence of butyrate or pyruvate, suggesting that the transporter may function as a tumor suppressor. Tumor cells meet their amino acid demands by inducing xCT/4F2hc, LAT1/4F2hc, ASCT2, and ATB0,+. xCT/4F2hc is related primarily to glutathione status, protection against oxidative stress, and cell cycle progression, whereas the other three transporters are related to amino acid nutrition. Pharmacologic blockade of LAT1/4F2hc, xCT/4F2hc, or ATB0,+ leads to inhibition of cancer cell growth. Since tumor cells selectively regulate these nutrient transporters to support their rapid growth, these transporters have potential as drug targets for cancer therapy.

Non-involvement of the human monocarboxylic acid transporter 1 (MCT1) in the transport of phenolic acid.

Phenolic acids such as p-coumaric acid and microbial metabolites of poorly absorbed polyphenols are absorbed by the monocarboxylic acid transporter (MCT)-mediated transport system which is identical to the fluorescein/H(+) cotransport system. We focus here on the physiological impact of MCT-mediated absorption and distribution. We examined whether MCT1, the best-characterized isoform found in almost all tissues, is involved in this MCT-mediated transport system. The induction of MCT1 expression in Caco-2 cells by a treatment with sodium butyrate (NaBut) did not increase the fluorescein permeability. Moreover, the transfection of Caco-2 cells with an expression vector encoding MCT1 caused no increase in either the permeability or uptake of fluorescein. Furthermore, in the MCT1-expressing oocytes, no increase of p-coumaric acid uptake was apparent, whereas the uptake of salicylic acid, a substrate of MCT1, nearly doubled. Our data therefore establish that MCT1 was not involved in the MCT-mediated transport of phenolic acids.

Food-drug interaction between ferulic acid and nateglinide involving the fluorescein/H+ cotransport system.

In clinical, patients usually take many kinds of drugs at the same time. Thus, drug-drug interactions involving transporters can often directly affect the therapeutic safety and efficacy of many drugs. However, there have been few studies on food-drug interactions involving transporters. Dietary polyphenols have been widely assumed to be beneficial to human health. Polyphenols are commercially prepared and used as functional foods. We report here for the first time that ferulic acid, which is widely used as a functional food, affects the transport of clinical agents. It is important to be aware of the potential of food-drug interactions and to act in order to prevent undesirable and harmful clinical consequences.