An Inhibitory Role for Sema4A in Antigen-Specific Allergic Asthma

Purpose

The class IV semaphorin Sema4A is critical for efficient T_H1 differentiation and Sema4a ^?/? mice exhibit impaired T_H1 immune responses. However, the role of Sema4A in T_H2 cell-mediated allergic diseases has not been fully studied. The aim of this study was to clarify the regulatory role possessed by Sema4A in mouse models of allergic diseases, particularly allergic asthma.

Methods

Sema4a ^?/? mice on a BALB/c background were examined for the development of allergic diseases. To induce experimental asthma, mice were sensitized with ovalbumin (OVA) followed by intranasal challenges with OVA. After challenge, airway hyperreactivity (AHR) and airway inflammation were evaluated. The role of Sema4A in asthma was examined using Sema4a ^?/? mice and Sema4A-Fc fusion proteins. The direct effects of Sema4A-Fc on antigen-specific effector CD4⁺ T cells were also examined.

Results

A fraction of Sema4a ^?/? BALB/c mice spontaneously developed skin lesions that resembled atopic dermatitis (AD) in humans. Furthermore, AHR, airway inflammation, and T_H2-type immune responses were enhanced in Sema4a ^?/? mice compared to wild type (WT) mice when immunized and challenged with OVA. In vivo systemic administration of Sema4A-Fc during the challenge period ameliorated AHR and lung inflammation and reduced the production of T_H2-type cytokines in WT mice. The inhibitory effects of Sema4A on airway inflammation were also observed in mice deficient in Tim-2, a Sema4A receptor. Finally, we showed that Sema4A-Fc directly inhibited IL-4-producing OVA-specific CD4⁺ T cells.

Conclusion

These results demonstrate that Sema4A plays an inhibitory role in T_H2-type allergic diseases, such as allergic asthma.     

Leukodystrophy caused by plasmalogen deficiency rescued by glyceryl 1‐myristyl ether treatment

Plasmalogens are the most abundant form of ether phospholipids in myelin and their deficiency causes Rhizomelic Chondrodysplasia Punctata (RCDP), a severe developmental disorder. Using the Gnpat‐knockout (KO) mouse as a model of RCDP, we determined the consequences of a plasmalogen deficiency during myelination and myelin homeostasis in the central nervous system (CNS). We unraveled that the lack of plasmalogens causes a generalized hypomyelination in several CNS regions including the optic nerve, corpus callosum and spinal cord. The defect in myelin content evolved to a progressive demyelination concomitant with generalized astrocytosis and white matter‐selective microgliosis. Oligodendrocyte precursor cells (OPC) and mature oligodendrocytes were abundant in the CNS of Gnpat KO mice during the active period of demyelination. Axonal loss was minimal in plasmalogen‐deficient mice, although axonal damage was observed in spinal cords from aged Gnpat KO mice. Characterization of the plasmalogen‐deficient myelin identified myelin basic protein and septin 7 as early markers of dysmyelination, whereas myelin‐associated glycoprotein was associated with the active demyelination phase. Using in vitro myelination assays, we unraveled that the intrinsic capacity of oligodendrocytes to ensheath and initiate membrane wrapping requires plasmalogens. The defect in plasmalogens was rescued with glyceryl 1‐myristyl ether [1‐O‐tetradecyl glycerol (1‐O‐TDG)], a novel alternative precursor in the plasmalogen biosynthesis pathway. 1‐O‐TDG treatment rescued myelination in plasmalogen‐deficient oligodendrocytes and in mutant mice. Our results demonstrate the importance of plasmalogens for oligodendrocyte function and myelin assembly, and identified a novel strategy to promote myelination in nervous tissue.     

Brain inflammation is accompanied by peripheral inflammation in <Emphasis Type="Italic">Cstb</Emphasis><Superscript><Emphasis Type="Italic">−/−</Emphasis></Superscript> mice,a model for progressive myoclonus epilepsy

Progressive myoclonus epilepsy of Unverricht-Lundborg type (EPM1) is an autosomal recessively inherited childhood-onset neurodegenerative disorder, characterized by myoclonus, seizures, and ataxia. Mutations in the cystatin B gene (CSTB) underlie EPM1. The CSTB-deficient (Cstb ^?/? ) mouse model recapitulates key features of EPM1, including myoclonic seizures. The mice show early microglial activation that precedes seizure onset and neuronal loss and leads to neuroinflammation. We here characterized the inflammatory phenotype of Cstb ^?/? mice in more detail. We found higher concentrations of chemokines and pro-inflammatory cytokines in the serum of Cstb ^?/? mice and higher CXCL13 expression in activated microglia in Cstb ^?/? compared to control mouse brains. The elevated chemokine levels were not accompanied by blood-brain barrier disruption, despite increased brain vascularization. Macrophages in the spleen and brain of Cstb ^?/? mice were predominantly pro-inflammatory. Taken together, these data show that CXCL13 expression is a hallmark of microglial activation in Cstb ^?/? mice and that the brain inflammation is linked to peripheral inflammatory changes, which might contribute to the disease pathology of EPM1.     

Interleukin-4 deficiency protects mice from acetaminophen-induced liver injury and inflammation by prevention of glutathione depletion

Objective

Interleukin-4 (IL-4) is a multifunctional cytokine involved in many diseases such as autoimmune hepatitis and idiosyncratic drug reactions. However, its role in acetaminophen (APAP)-induced liver injury remains unclear. Our objective was to evaluate the contribution of IL-4 to the pathogenesis of APAP-induced liver injury.

Methods

Balb/C (WT) and IL-4 knockout (IL-4^?/?) mice were orally overdosed with APAP. After 24 h, survival percentage, biochemical and morphological markers of liver injury, and tissue inflammation were assessed.

Results

IL-4^?/? mice were protected from APAP toxicity. Intravital confocal microscopy, tissue histology and serum ALT levels showed significantly less liver injury and inflammation than in the WT group, which may explain the increased survival rate of IL-4^?/? mice. In addition, IL-4^?/? mice had decreased production of tumor necrosis factor α, CXCL1 and interleukin-1β in the liver, but not in a remote site such as the lungs. Hepatic macrophage activation was markedly reduced in IL-4-deficient mice. In addition, glutathione depletion—a primary cause of APAP-mediated injury—was significantly attenuated in IL-4^?/? mice.

Conclusions

Taken together, our data demonstrate that IL-4^?/? mice are protected from APAP-induced liver injury due to reduced depletion of glutathione, which prevented liver damage and tissue inflammation.     

Repeated administration of the noradrenergic neurotoxin N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) modulates neuroinflammation and amyloid plaque load in mice bearing amyloid precursor protein and presenilin-1 mutant transgenes

Background

Data indicates anti-oxidant, anti-inflammatory and pro-cognitive properties of noradrenaline and analyses of post-mortem brain of Alzheimer's disease (AD) patients reveal major neuronal loss in the noradrenergic locus coeruleus (LC), the main source of CNS noradrenaline (NA). The LC has projections to brain regions vulnerable to amyloid deposition and lack of LC derived NA could play a role in the progression of neuroinflammation in AD. Previous studies reveal that intraperitoneal (IP) injection of the noradrenergic neurotoxin N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) can modulate neuroinflammation in amyloid over-expressing mice and in one study, DSP-4 exacerbated existing neurodegeneration.

Methods

TASTPM mice over-express human APP and beta amyloid protein and show age related cognitive decline and neuroinflammation. In the present studies, 5 month old C57/BL6 and TASTPM mice were injected once monthly for 6 months with a low dose of DSP-4 (5 mg kg^-1) or vehicle. At 8 and 11 months of age, mice were tested for cognitive ability and brains were examined for amyloid load and neuroinflammation.

Results

At 8 months of age there was no difference in LC tyrosine hydroxylase (TH) across all groups and cortical NA levels of TASTPM/DSP-4, WT/Vehicle and WT/DSP-4 were similar. NA levels were lowest in TASTPM/Vehicle. Messenger ribonucleic acid (mRNA) for various inflammatory markers were significantly increased in TASTPM/Vehicle compared with WT/Vehicle and by 8 months of age DSP-4 treatment modified this by reducing the levels of some of these markers in TASTPM. TASTPM/Vehicle showed increased astrocytosis and a significantly larger area of cortical amyloid plaque compared with TASTPM/DSP-4. However, by 11 months, NA levels were lowest in TASTPM/DSP-4 and there was a significant reduction in LC TH of TASTPM/DSP-4 only. Both TASTPM groups had comparable levels of amyloid, microglial activation and astrocytosis and mRNA for inflammatory markers was similar except for interleukin-1 beta which was increased by DSP-4. TASTPM mice were cognitively impaired at 8 and 11 months but DSP-4 did not modify this.

Conclusion

These data reveal that a low dose of DSP-4 can have varied effects on the modulation of amyloid plaque deposition and neuroinflammation in TASTPM mice dependent on the duration of dosing.     

Decreased arthritis severity in cathepsin L-deficient mice is attributed to an impaired T helper cell compartment

Objective

Cathepsin L (CL) is potentially involved in joint destruction and in antigen presentation in rheumatoid arthritis. In order to define the roles of this protease in arthritis development we analysed the antigen-induced arthritis (AIA) in CL-deficient (CL^?/?) mice.

Methods

Antigen-induced arthritis was induced in CL^?/? and wild-type mice. Complete CL deficiency resulted in an impaired positive selection of conventional CD4⁺ T helper (Th) cells and finally in a reduced number of Th cells. Thus, we addressed the effect of this phenotype by rescuing CD4⁺ Th cell numbers by transgenic expression of the human CL-like protease cathepsin V (hCV) in thymic epithelium of CL^?/? mice [Tg(K14-hCV);CL^?/?]. The arthritis development was monitored by measuring joint swelling. Joint inflammation and destruction were assessed histopathologically.

Results

The severity of AIA was decreased in CL^?/? mice characterized by reduced swelling, decreased inflammation and destruction, and diminished cellular and humoral immune responsiveness. AIA in Tg(K14-hCV);CL^?/? mice was associated with a reconstitution of all parameters by normalization of the ratio of regulatory to conventional T cells.

Conclusions

Cathepsin L has a significant impact on AIA severity by influencing the selection of Th cell populations in the thymus, but seems not play any significant role in the direct joint destruction.     

Liver X receptor activation attenuates plaque formation and improves vasomotor function of the aortic artery in atherosclerotic ApoE?/? mice

Aim

The severity of atherosclerosis is primarily determined by overall lipid metabolism and the degree of inflammation present within the vessel wall. We evaluated the effects of T-0901317, a liver X receptor agonist, on the atherosclerosis process, and especially on the endothelial function in ApoE^?/? mice.

Methods and results

ApoE^?/? mice were treated with LXR agonist T-0901317 (1?μmol/L) for 6?weeks. ApoE^?/? mice receiving T-0901317 were found to have markedly improved overall serum lipid profiles, albeit increased serum triglycerides. MRI imaging demonstrated that T-0901317 attenuated the atherosclerotic plaque burden in the aorta of ApoE^?/? mice. Transmission electron microscopy and immunohistochemistry revealed attenuated ultrastructural changes as well as enhanced expression of the ATP-binding cassette transporter ABCA1. In addition, treatment with the LXR agonist improved the vasomotor function of atherosclerotic arteries, as assessed by KCl/norepinephrine-induced vasoconstrictive and acetylcholine-induced vasorelaxation functional assays. In vitro studies showed increased ABCG1, phospho-Akt and phospho-eNOS expression in ApoE^?/? mice aorta endothelial cells (ECs) after T0901317 treatment.

Conclusion

The present study suggest that LXR agonists protect the endothelium against atherosclerotic insults by increasing ABCA1 and ABCG1 expression, and improve the endothelial-dependent vasomotor function probably by promoting Akt and eNOS phosphorylation.     

Chemokine CXCL13 activates p38 MAPK in the trigeminal ganglion after infraorbital nerve injury

Recent data demonstrated that chemokine CXCL13 mediates neuroinflammation and contributes to the maintenance of neuropathic pain after nerve injury in the spinal cord. Pro-nociceptive chemokines activate mitogen-activated protein kinases (MAPKs) which are potential signaling pathways contributing to the nociceptive behavior in inflammatory or neuropathic pain. However, whether activation of p38 and JNK MAPK signaling pathway in the trigeminal ganglion (TG) are involved in CXCL13 and its receptor CXCR5-mediated orofacial pain has not yet been clarified. Here, we show that the unilateral partial infraorbital nerve ligation (pIONL) induced a profound orofacial pain in wild-type (WT) mice. Western blot results showed that pIONL induced p38 but not JNK activation in the TG of WT mice. However, the orofacial pain induced by pIONL was alleviated in Cxcr5 ^?/? mice, and the activation of p38 was also abrogated in Cxcr5 ^?/? mice. Furthermore, intra-TG injection of CXCL13 evoked mechanical hypersensitivity and increased p-p38 expression in WT mice. But CXCL13 had no effect on pain behavior or p-p38 expression in Cxcr5 ^?/? mice. Finally, pretreatment with p38 inhibitor, SB203580, attenuated the pIONL-induced mechanical allodynia and decreased the mRNA expression of pro-inflammatory cytokines including tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) in the TG. Taken together, our data suggest that CXCL13 acts on CXCR5 to increase p38 activation and further contributes to the pathogenesis of orofacial neuropathic pain.     

Interleukin-33 amplifies IgE synthesis and triggers mast cell degranulation via interleukin-4 in naïve mice

Background

The regulation and function of IgE in healthy individuals and in antigen‐naïve animals is not well understood. IL‐33 administration increases serum IgE in mice with unknown mechanism. We tested the hypothesis that IL‐33 provides an antigen‐independent stimulus for IgE production and mast cell degranulation.

Methods

IL‐33 was administered to naïve wild‐type (WT), nude and ST2^?/?, IL‐4^?/?, IL4Rα^?/? and T‐or B‐cell‐specific IL‐4Rα^?/? mice. IgEand cytokines were quantified by ELISA. T‐ and B‐lymphocyte numbers and CD40L expression were determined by flow cytometry. Anaphylaxis was measured by temperature, mast cell degranulation and histamine release.

Results

IL‐33 enhanced IgE production in naïve WT, T‐IL‐4Rα^?/? but not in ST2^?/?, IL‐4^?/?, IL‐4Rα^?/? or B‐cell‐specific IL‐4Rα^?/? mice, demonstrating IL‐33 specificity and IL‐4 dependency. Moreover, IL‐4 was required for IL‐33‐induced B‐cell proliferation and T‐cell CD40L expression, which promotes IgE production. IL‐33‐induced IL‐4 production was mainly from innate cells including mast cells and eosinophils. IL‐33 increased mast cell surface IgE and triggered degranulation and systemic anaphylaxis in allergen‐naïve WT but not in IL‐4Rα^?/? mice.

Conclusion

IL‐33 amplifies IgE synthesis and triggers anaphylaxis in naïve mice via IL‐4, independent of allergen. IL‐33 may play an important role in nonatopic allergy and idiopathic anaphylaxis.

     

γδT cells but not αβT cells contribute to sepsis-induced white matter injury and motor abnormalities in mice

Background

Infection and sepsis are associated with brain white matter injury in preterm infants and the subsequent development of cerebral palsy.

Methods

In the present study, we used a neonatal mouse sepsis-induced white matter injury model to determine the contribution of different T cell subsets (αβT cells and γδT cells) to white matter injury and consequent behavioral changes. C57BL/6J wild-type (WT), T cell receptor (TCR) δ-deficient (Tcrd ^?/?, lacking γδT cells), and TCRα-deficient (Tcra ^?/?, lacking αβT cells) mice were administered with lipopolysaccharide (LPS) at postnatal day (PND) 2. Brain myelination was examined at PNDs 12, 26, and 60. Motor function and anxiety-like behavior were evaluated at PND 26 or 30 using DigiGait analysis and an elevated plus maze.

Results

White matter development was normal in Tcrd ^?/? and Tcrα ^?/? compared to WT mice. LPS exposure induced reductions in white matter tissue volume in WT and Tcrα ^?/? mice, but not in the Tcrd ^?/? mice, compared with the saline-treated groups. Neither LPS administration nor the T cell deficiency affected anxiety behavior in these mice as determined with the elevated plus maze. DigiGait analysis revealed motor function deficiency after LPS-induced sepsis in both WT and Tcrα ^?/? mice, but no such effect was observed in Tcrd ^?/? mice.

Conclusions

Our results suggest that γδT cells but not αβT cells contribute to sepsis-induced white matter injury and subsequent motor function abnormalities in early life. Modulating the activity of γδT cells in the early stages of preterm white matter injury might represent a novel therapeutic strategy for the treatment of perinatal brain injury.

     

A lack of association between hyperserotonemia and the increased frequency of serum anti-myelin basic protein auto-antibodies in autistic children

Background

The purpose of this study was to investigate whether localized peripheral inflammation, such as osteoarthritis, contributes to neuroinflammation and neurodegenerative disease in vivo.

Methods

We employed the inducible Col1-IL1β^XAT mouse model of osteoarthritis, in which induction of osteoarthritis in the knees and temporomandibular joints resulted in astrocyte and microglial activation in the brain, accompanied by upregulation of inflammation-related gene expression. The biological significance of the link between peripheral and brain inflammation was explored in the APP/PS1 mouse model of Alzheimer's disease (AD) whereby osteoarthritis resulted in neuroinflammation as well as exacerbation and acceleration of AD pathology.

Results

Induction of osteoarthritis exacerbated and accelerated the development of neuroinflammation, as assessed by glial cell activation and quantification of inflammation-related mRNAs, as well as Aβ pathology, assessed by the number and size of amyloid plaques, in the APP/PS1; Col1-IL1β^XAT compound transgenic mouse.

Conclusion

This work supports a model by which peripheral inflammation triggers the development of neuroinflammation and subsequently the induction of AD pathology. Better understanding of the link between peripheral localized inflammation, whether in the form of osteoarthritis, atherosclerosis or other conditions, and brain inflammation, may prove critical to our understanding of the pathophysiology of disorders such as Alzheimer's, Parkinson's and other neurodegenerative diseases.     

Appearance of claudin-5<Superscript>+</Superscript> leukocytes in the central nervous system during neuroinflammation: a novel role for endothelial-derived extracellular vesicles

Background

The mechanism of leukocyte transendothelial migration (TEM) across the highly restrictive blood-brain barrier (BBB) remains enigmatic, with paracellular TEM thought to require leukocytes to somehow navigate the obstructive endothelial tight junctions (TJs). Transient interactions between TJ proteins on the respective leukocyte and endothelial surfaces have been proposed as one mechanism for TEM. Given the expanding role of extracellular vesicles (EVs) in intercellular communication, we investigated whether EVs derived from brain microvascular endothelial cells (BMEC) of the BBB may play a role in transferring a major TJ protein, claudin-5 (CLN-5), to leukocytes as a possible basis for such a mechanism during neuroinflammation.

Methods

High-resolution 3D confocal imaging was used to highlight CLN-5 immunoreactivity in the central nervous system (CNS) and on leukocytes of mice with the neuroinflammatory condition experimental autoimmune encephalomyelitis (EAE). Both Western blotting of circulating leukocytes from wild-type mice and fluorescence imaging of leukocyte-associated eGFP-CLN-5 in the blood and CNS of endothelial-targeted, Tie-2-eGFP-CLN-5 transgenic mice were used to confirm the presence of CLN-5 protein on these cells. EVs were isolated from TNF-α-stimulated BMEC cultures and blood plasma of Tie-2-eGFP-CLN-5 mice with EAE and evaluated for CLN-5 protein by Western blotting and fluorescence-activated cell sorting (FACS), respectively. Confocal imaging and FACS were used to detect binding of endothelial-derived EVs from these two sources to leukocytes in vitro. Serial electron microscopy (serial EM) and 3D contour-based surface reconstruction were employed to view EV-like structures at the leukocyte:BBB interface in situ in inflamed CNS microvessels.

Results

A subpopulation of leukocytes immunoreactive for CLN-5 on their surface was seen to infiltrate the CNS of mice with EAE and reside in close apposition to inflamed vessels. Confocal imaging of immunostained samples and Western blotting established the presence of CLN-5⁺ leukocytes in blood as well, implying these cells are present prior to TEM. Moreover, imaging of inflamed CNS vessels and the associated perivascular cell infiltrates from Tie-2-eGFP-CLN-5 mice with EAE revealed leukocytes bearing the eGFP label, further supporting the hypothesis CLN-5 is transferred from endothelial cells to circulating leukocytes in vivo. Western blotting of BMEC-derived EVs, corresponding in size to both exosomes and microvesicles, and FACS analysis of plasma-derived EVs from Tie-2-eGFP-CLN-5 mice with EAE validated expression of CLN-5 by EVs of endothelial origin. Confocal imaging and FACS further revealed both PKH-67-labeled EVs from cultured BMECs and eGFP-CLN-5⁺ EVs from plasma of Tie-2-eGFP-CLN-5 mice with EAE can bind to leukocytes. Lastly, serial EM and 3D contour-based surface reconstruction revealed a close association of EV-like structures between the marginating leukocytes and BMECs in situ during EAE.

Conclusions

During neuroinflammation, CLN-5⁺ leukocytes appear in the CNS, and both CLN-5⁺ leukocytes and CLN-5⁺ EVs are detected in the blood. As endothelial cells transfer CLN-5⁺ to leukocytes in vivo, and EVs released from BMEC bind to leukocytes in vitro, EVs may serve as the vehicles to transfer CLN-5 protein at sites of leukocyte:endothelial contact along the BBB. This action may be a prelude to facilitate TEM through the formation of temporary TJ protein bridges between these two cell types.

     

Mitochondrial aldehyde dehydrogenase (ALDH2) protects against streptozotocin-induced diabetic cardiomyopathy: role of GSK3β and mitochondrial function

Background

Mitochondrial aldehyde dehydrogenase (ALDH2) displays some promise in the protection against cardiovascular diseases although its role in diabetes has not been elucidated.

Methods

This study was designed to evaluate the impact of ALDH2 on streptozotocin-induced diabetic cardiomyopathy. Friendly virus B(FVB) and ALDH2 transgenic mice were treated with streptozotocin (intraperitoneal injection of 200 mg/kg) to induce diabetes.

Results

Echocardiographic evaluation revealed reduced fractional shortening, increased end-systolic and -diastolic diameter, and decreased wall thickness in streptozotocin-treated FVB mice. Streptozotocin led to a reduced respiratory exchange ratio; myocardial apoptosis and mitochondrial damage; cardiomyocyte contractile and intracellular Ca²⁺ defects, including depressed peak shortening and maximal velocity of shortening and relengthening; prolonged duration of shortening and relengthening; and dampened intracellular Ca²⁺ rise and clearance. Western blot analysis revealed disrupted phosphorylation of Akt, glycogen synthase kinase-3?? and Foxo3a (but not mammalian target of rapamycin), elevated PTEN phosphorylation and downregulated expression of mitochondrial proteins, peroxisome proliferator-activated receptor ?? coactivator 1?? and UCP-2. Intriguingly, ALDH2 attenuated or ablated streptozotocin-induced echocardiographic, mitochondrial, apoptotic and myocardial contractile and intracellular Ca²⁺ anomalies as well as changes in the phosphorylation of Akt, glycogen synthase kinase-3??, Foxo3a and phosphatase and tensin homologue on chromosome ten, despite persistent hyperglycemia and a low respiratory exchange ratio. In vitro data revealed that the ALDH2 activator Alda-1 and glycogen synthase kinase-3?? inhibition protected against high glucose-induced mitochondrial and mechanical anomalies, the effect of which was cancelled by mitochondrial uncoupling.

Conclusions

In summary, our data revealed that ALDH2 acted against diabetes-induced cardiac contractile and intracellular Ca²⁺ dysregulation, possibly through regulation of apoptosis, glycogen synthase kinase-3?? activation and mitochondrial function independent of the global metabolic profile.     

A mouse model of sitosterolemia: absence of Abcg8/sterolin-2 results in failure to secrete biliary cholesterol

Background

Mutations in either of two genes comprising the STSL locus, ATP-binding cassette (ABC)-transporters ABCG5 (encoding sterolin-1) and ABCG8 (encoding sterolin-2), result in sitosterolemia, a rare autosomal recessive disorder of sterol trafficking characterized by increased plasma plant sterol levels. Based upon the genetics of sitosterolemia, ABCG5/sterolin-1 and ABCG8/sterolin-2 are hypothesized to function as obligate heterodimers. No phenotypic difference has yet been described in humans with complete defects in either ABCG5 or ABCG8. These proteins, based upon the defects in humans, are responsible for regulating dietary sterol entry and biliary sterol secretion.

Methods

In order to mimic the human disease, we created, by a targeted disruption, a mouse model of sitosterolemia resulting in Abcg8/sterolin-2 deficiency alone. Homozygous knockout mice are viable and exhibit sitosterolemia.

Results

Mice deficient in Abcg8 have significantly increased plasma and tissue plant sterol levels (sitosterol and campesterol) consistent with sitosterolemia. Interestingly, Abcg5/sterolin-1 was expressed in both liver and intestine in Abcg8/sterolin-2 deficient mice and continued to show an apical expression. Remarkably, Abcg8 deficient mice had an impaired ability to secrete cholesterol into bile, but still maintained the ability to secrete sitosterol. We also report an intermediate phenotype in the heterozygous Abcg8+/- mice that are not sitosterolemic, but have a decreased level of biliary sterol secretion relative to wild-type mice.

Conclusion

These data indicate that Abcg8/sterolin-2 is necessary for biliary sterol secretion and that loss of Abcg8/sterolin-2 has a more profound effect upon biliary cholesterol secretion than sitosterol. Since biliary sitosterol secretion is preserved, although not elevated in the sitosterolemic mice, this observation suggests that mechanisms other than by Abcg8/sterolin-2 may be responsible for its secretion into bile.     

The phosphate transporter NaPi-IIa determines the rapid renal adaptation to dietary phosphate intake in mouse irrespective of persistently high FGF23 levels

Renal reabsorption of inorganic phosphate (Pi) is mediated by the phosphate transporters NaPi-IIa, NaPi-IIc, and Pit-2 in the proximal tubule brush border membrane (BBM). Dietary Pi intake regulates these transporters; however, the contribution of the specific isoforms to the rapid and slow phase is not fully clarified. Moreover, the regulation of PTH and FGF23, two major phosphaturic hormones, during the adaptive phase has not been correlated. C57/BL6 and NaPi-IIa^?/? mice received 5 days either 1.2 % (HPD) or 0.1 % (LPD) Pi-containing diets. Thereafter, some mice were acutely switched to LPD or HPD. Plasma Pi concentrations were similar under chronic diets, but lower when mice were acutely switched to LPD. Urinary Pi excretion was similar in C57/BL6 and NaPi-IIa^?/? mice under HPD. During chronic LPD, NaPi-IIa^?/? mice lost phosphate in urine compensated by higher intestinal Pi absorption. During the acute HPD-to-LPD switch, NaPi-IIa^?/? mice exhibited a delayed decrease in urinary Pi excretion. PTH was acutely regulated by low dietary Pi intake. FGF23 did not respond to low Pi intake within 8 h whereas the phospho-adaptator protein FRS2α necessary for FGF-receptor cell signaling was downregulated. BBM Pi transport activity and NaPi-IIa but not NaPi-IIc and Pit-2 abundance acutely adapted to diets in C57/BL6 mice. In NaPi-IIa^?/?, Pi transport activity was low and did not adapt. Thus, NaPi-IIa mediates the fast adaptation to Pi intake and is upregulated during the adaptation to low Pi despite persistently high FGF23 levels. The sensitivity to FGF23 may be regulated by adapting FRS2α abundance and phosphorylation.