Inhibition of the lactic acid transporters MCT1 and MCT4 as an underlying mechanism for drug-induced myopathy

Les myopathies induites par les médicaments représentent un effet secondaire sérieux causé par plusieurs médicaments. Ces symptômes musculaires varient de myalgies légères avec ou sans élévation de créatine kinase, faiblesse musculaire, myosite, jusqu’à de rares rhabdomyolyses potentiellement mortelles. Bien que les myalgies légères soient tolérables, les myopathies chroniques affectent la qualité de vie des patients, requérant souvent la cessation d’une thérapie efficace. Le mécanisme sous-jacent à ces myotoxicités causées par les médicaments est connu pour certains composés, mais demeure obscur pour plusieurs (ex. statines). Les statines constituent une thérapie efficace pour la diminution du cholestérol, mais elles sont reconnues pour causer ces effets secondaires. De nombreux facteurs augmentant les concentrations plasmatiques de statines (ex. doses élevées, interactions médicamenteuses, polymorphismes génétiques) semblent être liés à une fréquence de myotoxicité plus élevée. Conséquemment, le métabolisme et le transport des médicaments, contrôlant l’absorption globale, la distribution et l’élimination, peuvent devenir importants. Cependant, ces facteurs peuvent seulement expliquer partiellement les désordres musculaires observés. Même si plusieurs mécanismes sont proposés pour les myotoxicités induites par les statines, le mécanisme exact responsable de cet effet est controversé, puisque les études rapportent des résultats contradictoires. Puisque l’exercice semble exacerber les douleurs musculaires chez les patients prenant des statines, l’hypothèse derrière ce projet est que le transport de l’acide L-lactique par les transporteurs de monocarboxylates serait impliqué dans le développement des myotoxicités. Puisque l’acide L-lactique est l’un des sous-produits majeurs résultant de l’activité physique, son élimination efficace des cellules musculaires est essentielle. L’administration de médicaments inhibant compétitivement ces transporteurs pourrait mener à une perturbation de l’homéostasie de l’acide L-lactique et à des désordres musculaires. L’objectif du premier volet de cette étude était d’évaluer le potentiel d’inhibition des médicaments acides sur le transport d’acide L-lactique en utilisant les lignées cellulaires cancéreuses Hs578T et MDA-MB-231 exprimant sélectivement MCT1 ou MCT4, respectivement. Ces lignées cellulaires ont permis la caractérisation des transporteurs avec la détermination de leurs paramètres cinétiques et d’inhibition. Le but principal du deuxième volet de cette étude était de confirmer le potentiel d’inhibition de l’atorvastatine, simvastatine, rosuvastatine et loratadine sur le transport d’acide L-lactique dans un cadre plus physiologique en utilisant des cellules musculaires squelettiques primaires humaines (SkMC). L’objectif global de ce projet de doctorat était de mieux comprendre les mécanismes derrière certaines myopathies induites par les médicaments, plus spécifiquement celles induites par les statines et la loratadine, en étudiant les transporteurs de monocarboxylates impliqués dans le transport d’acide L-lactique et l’homéostasie du pH dans le muscle. La loratadine et l’atorvastatine ont démontré le meilleur potentiel d’inhibition de l’efflux d’acide L-lactique dans les lignées cellulaires, une observation confirmée dans les SkMC. Cette inhibition pourrait causer une accumulation intracellulaire d’acide L-lactique menant à une acidification et à des désordres musculaires. De futures études dans des modèles in vivo sont requises pour confirmer l’impact physiologique de nos résultats dans un cadre clinique. Ces données permettraient une meilleure compréhension des myopathies induites par les statines et la loratadine et permettront ainsi de prévenir leur occurrence en optimisant les stratégies thérapeutiques.Drug-induced myopathy is a serious side effect caused by various widely-administered medications. These muscle-related symptoms range from mild myalgia with or without creatine kinase increase, muscle weakness, myositis, to rare life-threatening rhabdomyolysis. While mild myalgias can be tolerable, chronic myopathies can affect the patients' quality of life, frequently requiring the cessation of an effective drug. The underlying mechanism of these drug-induced myotoxicities is known for some drugs but remains unclear for most (e.g. statins). Statins constitute an effective cholesterol-lowering therapy, but they are known to cause these adverse drug reactions. Various factors increasing statin plasma levels (e.g. high doses, drug-drug interactions, genetic polymorphisms) seem to be linked with a higher occurrence of myotoxicity. Consequently, systemic drug metabolism and transport, controlling overall absorption, distribution and elimination, can become important. However, these factors only partly explain the observed muscular disorders. Although there are several proposed mechanisms for statin-induced myotoxicity, the exact mechanism responsible for this effect is still debated with studies reporting conflicting results. Since exercise seems to exacerbate muscle pain in patients under statin treatment, the premise of this project is that L-lactic acid transport via the monocarboxylate transporters is involved in the development of drug-induced myopathy. Since lactic acid is one of the major byproducts resulting from physical activity, its efficient removal from the muscle cells is essential. Therefore, the administration of drugs competitively inhibiting those transporters may potentially lead to perturbation of L-lactic acid homeostasis and muscular disorders. The aim of the first part of this study was to assess the inhibitory potential of acidic drugs on L-lactic acid transport using breast cancer cell lines Hs578T and MDA-MB-231, which selectively express MCT1 or MCT4, respectively. These cell lines allowed transporter characterization with the determination of their kinetic parameters and inhibition. The main objective of the second part of this study was to confirm the inhibitory potentials of atorvastatin, simvastatin, rosuvastatin and loratadine on L-lactic acid transport in a more physiological setting using primary human skeletal muscle cells (SkMC). The overall goal of this doctoral project was to better understand the mechanisms behind certain drug-induced myopathies, more specifically those induced by statins and loratadine, by studying monocarboxylate transporters involved in lactic acid transport and pH homeostasis in the muscle. Loratadine and atorvastatin demonstrated the greatest potency for inhibition of L-lactic acid efflux first in cancer cell lines, an observation confirmed in SkMC. This inhibition may cause an accumulation of intracellular L-lactic acid leading to acidification and muscular disorders. Further studies with in vivo models are required to confirm the physiological impact of our findings in a clinical setting. These data will help understand statin- and loratadine-induced myopathy and prevent its occurrence by optimizing treatment strategies

Interrater and intrarater reliability in rating velopharyngeal gap size

Also available in print.A dissertation submitted in partial fulfilment of the requirements for the Bachelor of Science (Speech and Hearing Sciences), The University of Hong Kong, June 30, 2007.Thesis (B.Sc)--University of Hong Kong, 2007.published_or_final_versionSpeech and Hearing SciencesBachelorBachelor of Science in Speech and Hearing Science

Distortion product otoacoustic emissions in patients with haemophilia

Includes bibliographical references (p. 22-24)."A dissertation submitted in partial fulfillment of the requirements for the Bachelor of Science (Speech and Hearing Sciences), The University of Hong Kong, June 30, 2010."Thesis (B.Sc)--University of Hong Kong, 2010.This study investigated the cochlear function of 13 haemophilic patients using distortion product otoacoustic emission (DPOAE) assessment techniques. Out of the 26 ears examined, a minority of them produced responses that were considered to be impaired. A positive correlation was found between clotting factor level and DPOAE amplitude, and DPOAE/ noise ratio at high frequencies (4, 6, 8 kHz). Clotting factor level, which is indicative of the severity of haemophilia, may be related to hearing status. It was also revealed that DPOAE amplitude and DPOAE/ noise ratio at low frequencies (1, 2, 3 kHz) were greater than those at high frequencies (4, 6, 8 kHz). Therefore the high-frequency cochlear region may be more sensitive to damage. Intracranial micro-haemorrhage in haemophilic individuals may be linked to an increased risk of cochlear damage. Further studies may use brain computed tomography scan to explore the possible link between intracranial micro-haemorrhage and hearing loss.published_or_final_versionSpeech and Hearing SciencesBachelorBachelor of Science in Speech and Hearing Science

Transformation of Magnaporthe grisea to phosphinothricin resistance using the bar gene from Streptomyces hygroscopicus

Three transformation systems have been reported for the rice blast fungus Magnaporthe grisea (Parsons et al. 1987 Proc. Natl. Acad. Sci. USA 84:4161-4165; Daboussi et al. 1989 Curr. Genet. 15:453-456; Leung et al. 1990 Curr. Genet. 17:409-411). Among these three selection systems, only hygromycin B resistance provides a dominant selection that can be used for any wild type strain. A second dominant selection marker is needed to transform strains that are already hygromycin B resistant

Characterizing genetic diversity and creating novel gene pools in rice for trait dissection and gene function discovery

Rice diversity is the foundation for rice improvement programs. At IRRI, over 100,000 rice accessions are deposited, and intelligent use of this diversity can not only help solve current production problems but also create future production opportunities and tackle climate change challenges. To fully explore and utilize rice diversity, two ingredients are needed: 1 - the genetic blueprints of diverse rice accessions in use, 2 - plant populations with recombined genotypes allowing expression of phenotypic variation and discovery of new genes/QTLs for use in breeding programs. Sequencing of the genomes & obtaining SNP genotypes of many rice accessions is feasible due to decreasing cost of advanced DNA sequencing technologies. Coupled with the creation of populations suitable for trait dissection / phenotyping, discovery of gene functions and allelic variations causal to important agronomic traits becomes possible. This in turn will provide rich biological evidences to the rice/cereal crop genome annotation community

Loss of Yeast Peroxiredoxin Tsa1p Induces Genome Instability through Activation of the DNA Damage Checkpoint and Elevation of dNTP Levels

Peroxiredoxins are a family of antioxidant enzymes critically involved in cellular defense and signaling. Particularly, yeast peroxiredoxin Tsa1p is thought to play a role in the maintenance of genome integrity, but the underlying mechanism is not understood. In this study, we took a genetic approach to investigate the cause of genome instability in tsa1Δ cells. Strong genetic interactions of TSA1 with DNA damage checkpoint components DUN1, SML1, and CRT1 were found when mutant cells were analyzed for either sensitivity to DNA damage or rate of spontaneous base substitutions. An elevation in intracellular dNTP production was observed in tsa1Δ cells. This was associated with constitutive activation of the DNA damage checkpoint as indicated by phosphorylation of Rad9/Rad53p, reduced steady-state amount of Sml1p, and induction of RNR and HUG1 genes. In addition, defects in the DNA damage checkpoint did not modulate intracellular level of reactive oxygen species, but suppressed the mutator phenotype of tsa1Δ cells. On the contrary, overexpression of RNR1 exacerbated this phenotype by increasing dNTP levels. Taken together, our findings uncover a new role of TSA1 in preventing the overproduction of dNTPs, which is a root cause of genome instability