The mdx mouse as a model for carnitine deficiency in the pathogenesis of Duchenne muscular dystrophy.

INTRODUCTION: Muscle and cardiac metabolism are dependent on the oxidation of fats and glucose for adenosine triphosphate production, for which L-carnitine is an essential cofactor. METHODS: We measured muscle carnitine concentrations in skeletal muscles, diaphragm, and ventricles of C57BL/10ScSn-DMDmdx/J mice (n = 10) and compared them with wild-type C57BL/6J (n = 3), C57BL/10 (n = 10), and C3H (n = 12) mice. Citrate synthase (CS) activity was measured in quadriceps/gluteals and ventricles of mdx and wild-type mice. RESULTS: We found significantly lower tissue carnitine in quadriceps/gluteus (P < 0.05) and ventricle (P < 0.05), but not diaphragm of mdx mice, when compared with controls. CS activity was increased in mdx quadriceps/gluteus (P < 0.03) and ventricle (P < 0.02). This suggests compensatory mitochondrial biogenesis. CONCLUSIONS: Decreased tissue carnitine has implications for reduced fatty acid and glucose oxidation in mdx quadriceps/gluteus and ventricle. The mdx mouse may be a useful model for studying the role of muscle carnitine deficiency in DMD bioenergetic insufficiency and providing a targeted and timed rationale for L-carnitine therapy.

Upregulation of mammary gland OCTNs maintains carnitine homeostasis in suckling infants.

BACKGROUND: Transport of L-carnitine, essential cofactor of fatty acid metabolism, into breast milk is critical for the normal growth and development of the suckling infant. OBJECTIVE: To increase understanding of developmental expression of carnitine/organic cation (Octn) transporter family at different stages of murine breast development for carnitine delivery. METHODS: We applied our transporter-specific antibodies to mOctn1, mOctn2 and mOctn3 to sections of mammary glands of virginal non-lactating, pregnant, late lactating and post-lactating C3H females. RESULTS: We demonstrated differential expression of mOctn1, -2 and -3 in epithelial ducts, specialized myoepithelial cells and fatty stroma. There was notable upregulation of all three Octns and mRNA by RT-PCR concurrent with an increase in epithelial ducts in breasts of pregnant (15days gestation) and lactating mice (15-days post-partum) compared to virginal 6 week old females, and notable downregulation in expression of Octns 15 days after cessation of lactation. In lactating murine mammary gland at 15 days post-partum, there was a marked increase of fat globules in epithelial ducts. Octn1 and Octn2 had similar expression patterns in lactating gland cells which formed fat globules that were exocytosed into the lumen of alveoli along with transporters Octn1 and Octn2. Octn3 was primarily localized to myoepithelial cells surrounding the ducts at all stages of breast development. CONCLUSIONS: There is a dynamic upregulation of the Octn family in pregnant and lactating breasts which likely provides the suckling infant with adequate carnitine for the rapid postnatal upregulation of fatty acid oxidation and ketogenesis critical for cerebral energy metabolism during fasting hypoglycemia.

Expression patterns of the organic cation/carnitine transporter family in adult murine brain.

UNLABELLED: Organic cation/carnitine transporters transport carnitine, drugs, and xenobiotics (e.g. choline, acetylcarnitine, betaine, valproic acid), and are expressed in muscle, heart, blood vessels, kidney, gut, etc. OBJECTIVE: To characterize expression patterns of mOctn1, -2 and -3 in murine brain. METHODS: We applied our transporter-specific antibodies to mOctn1, -2 and -3, followed by 2 0 antibody and DAB peroxidase detection to serial adult murine brain sections counterstained with hematoxylin. RESULTS: All three transporters showed strong expression in the external plexiform layer of the olfactory bulb and in olfactory nerve, the molecular layer and neuronal processes of input fibres extending vertically in motor cortex, in the dendritic arborization of the cornu ammonis and dendate gyrus (hippocampus), neuronal processes in the arcuate nucleus (hypothalamus), choroid plexus cells, and neuronal cell bodies and dendrites of cranial nerve nuclei V and VII. In the cerebellum, all three transporters were strongly expressed in dendritic processes of Purkinje cells, but Octn1 and -2 were expressed more strongly than Octn3 in Purkinje cell bodies. In spinal cord, Octn1, -2 and -3 were prominent in axons and dendritic end-arborizations of spinal cord neurons in both ascending and descending white matter tracts, whereas Octn3 was also strongly expressed in grey matter, specifically in anterior horn cell bodies. Octn3 was weakly expressed in glomerular layer neuronal cell bodies of olfactory bulb. CONCLUSIONS: hOCTN2 deficiency presents with carnitine-responsive cardiomyopathy, myopathy and hypoglycemic, hypoketotic coma with strokes, seizures and delays. In mouse, Octn1, -2 and -3 are expressed in many regions throughout the central nervous system with a pattern suggestive of roles in modulating cerebral bioenergetics and in acetylcholine generation for neurotransmission in olfactory, satiety, limbic, memory, motor and sensory functions. This distribution may play a role in the pattern of neurological injury that occurs in hOCTN2 deficiency during catabolic episodes of hypoglycemic, hypoketotic encephalopathy and which may manifest with cognitive impairment, hypotonia and seizures.

Acceptance of related and unrelated cardiac allografts in neonatally tolerized mice is cardio-specific and transferable by regulatory CD4+ T cells.

BACKGROUND: Non-donor-specific cardiac allograft acceptance is induced in C3H/He (C3H; H-2k) recipients injected as neonates with allogeneic BALB/c (BALB; H-2d) fetal liver cells (FLC). This occurs despite intact reactivity to donor-type and third-party alloantigens in in vitro assays and skin transplants. To investigate a role for regulatory T cells, we performed adoptive transfer studies and specifically assessed CD4+ and CD4- T cells. METHODS: Three cell populations (splenocytes, CD4+, CD4-) derived from neonatally-treated mice with accepted C57BL/6 (B6; H-2b) third-party cardiac grafts were adoptively transferred into sub-lethally-irradiated C3H mice. Reconstituted mice were challenged with B6 cardiac grafts, B6 skin grafts, or unrelated cardiac grafts. Separated cells were assessed in vitro. RESULTS: B6, BALB, and NZW (H-2z) graft acceptance was transferred by unfractionated splenocytes. CD4+ cells transferred B6 graft acceptance (85% survival > 100 days). CD4- cells, unfractionated cells from naive or only irradiated mice, and unfractionated cells from neonatally-treated non-transplanted C3H mice rejected grafts within 35 days. No inoculum induced skin graft acceptance. Co-cultured assays confirmed the suppressive function of CD4+ cells in vitro. CONCLUSIONS: Cardiac allograft acceptance in our model is regulated by CD4+ cells. The regulatory cell population is induced by the cardiac graft itself and mediates in vivo cardiac graft acceptance in a tissue-specific but not donor-strain-specific manner.

Organic cation/carnitine transporter family expression patterns in adult murine heart.

Organic cation/carnitine transporters transport carnitine, drugs, and xenobiotics (e.g. choline, acetylcarnitine, quinidine, and verapamil), and are expressed in muscle, heart, blood vessels, etc. To characterize expression patterns of mOctn1, -2, and -3 in adult murine heart, we applied our transporter-specific antibodies to mOctn1, -2, and -3, followed by secondary antibody and DAB peroxidase detection to adult murine heart sections counterstained with hematoxylin. All three transporters showed strong expression in cardiomyocytes, lamina fibrosa of cardiac valves, great arteries and intermuscular arterioles, and a striking differential expression in the vagal innervated sinoatrial and atrioventricular nodes. The hOCTN2 deficiency presents with carnitine-responsive cardiomyopathy. Octn1, -2, and -3 are expressed in many regions of murine heart with a pattern suggestive of potential roles in modulating myocardial bioenergetics, valvular function, and acetylcholine generation for parasympathetic vagal innervation of the cardiac conduction system. This distribution may play a role in the hypertrophic cardiomyopathy seen in hOCTN2 deficiency, and may also affect the absorption/elimination of organic cationic cardiac drugs.