Autistic disorder in patients with Williams-Beuren syndrome: a reconsideration of the Williams-Beuren syndrome phenotype

Background

Williams-Beuren syndrome (WBS), a rare developmental disorder caused by deletion of contiguous genes at 7q11.23, has been characterized by strengths in socialization (overfriendliness) and communication (excessive talkativeness). WBS has been often considered as the polar opposite behavioral phenotype to autism. Our objective was to better understand the range of phenotypic expression in WBS and the relationship between WBS and autistic disorder.

Methodology

The study was conducted on 9 French individuals aged from 4 to 37 years old with autistic disorder associated with WBS. Behavioral assessments were performed using Autism Diagnostic Interview-Revised (ADI-R) and Autism Diagnostic Observation Schedule (ADOS) scales. Molecular characterization of the WBS critical region was performed by FISH.

Findings

FISH analysis indicated that all 9 patients displayed the common WBS deletion. All 9 patients met ADI-R and ADOS diagnostic criteria for autism, displaying stereotypies and severe impairments in social interaction and communication (including the absence of expressive language). Additionally, patients showed improvement in social communication over time.

Conclusions

The results indicate that comorbid autism and WBS is more frequent than expected and suggest that the common WBS deletion can result in a continuum of social communication impairment, ranging from excessive talkativeness and overfriendliness to absence of verbal language and poor social relationships. Appreciation of the possible co-occurrence of WBS and autism challenges the common view that WBS represents the opposite behavioral phenotype of autism, and might lead to improved recognition of WBS in individuals diagnosed with autism.     

Rapamycin ameliorates dystrophic phenotype in mdx mouse skeletal muscle

Duchenne muscular dystrophy (DMD) is an X-linked, lethal, degenerative disease that results from mutations in the dystrophin gene, causing necrosis and inflammation in skeletal muscle tissue. Treatments that reduce muscle fiber destruction and immune cell infiltration can ameliorate DMD pathology. We treated the mdx mouse, a model for DMD, with the immunosuppressant drug rapamycin (RAPA) both locally and systemically to examine its effects on dystrophic mdx muscles. We observed a significant reduction of muscle fiber necrosis in treated mdx mouse tibialis anterior (TA) and diaphragm (Dia) muscles 6 wks post-treatment. This effect was associated with a significant reduction in infiltration of effector CD4(+) and CD8(+) T cells in skeletal muscle tissue, while Foxp3(+) regulatory T cells were preserved. Because RAPA exerts its effects through the mammalian target of RAPA (mTOR), we studied the activation of mTOR in mdx TA and Dia with and without RAPA treatment. Surprisingly, mTOR activation levels in mdx TA were not different from control C57BL/10 (B10). However, mTOR activation was different in Dia between mdx and B10; mTOR activation levels did not rise between 6 and 12 wks of age in mdx Dia muscle, whereas a rise in mTOR activation level was observed in B10 Dia muscle. Furthermore, mdx Dia, but not TA, muscle mTOR activation was responsive to RAPA treatment.     

Genotype, phenotype, and karyotype correlation in the XO mouse model of Turner Syndrome

The murine model for Turner Syndrome is the XO mouse. Unlike their human counterparts, XO mice are typically fertile, and their lack of a second sex chromosome can be transmitted from one generation to the next as an X-linked dominant trait with male lethality. The introduction of an X-linked coat-color marker (tabby) has greatly facilitated the maintenance of this useful mouse strain. XO mice can be produced in large numbers, generation after generation, and rapidly identified on the basis of their sex and coat color. Although this breeding scheme appears to be effective at the phenotype level, its utility has never been conclusively proved at the molecular or cytogenetic levels. Here, we clone and sequence the tabby deletion break point and present a multiplex polymerase chain reaction-based assay for the tabby mutation. By combining the results of this assay with whole-chromosome painting data, we demonstrate that genotype, phenotype, and karyotype all show perfect correlation in the publicly available XO breeding stock. This work lays the foundation for the use of this strain to study Turner Syndrome in particular and the X chromosome in general.     

Reduction in neuronal L-type calcium channel activity in a double knock-in mouse model of Alzheimer's disease

Increased function of neuronal L-type voltage-sensitive Ca^2 + channels (L-VSCCs) is strongly linked to impaired memory and altered hippocampal synaptic plasticity in aged rats. However, no studies have directly assessed L-VSCC function in any of the common mouse models of Alzheimer's disease where neurologic deficits are typically more robust. Here, we used cell-attached patch-clamp recording techniques to measure L-VSCC activity in CA1 pyramidal neurons of partially dissociated hippocampal “zipper” slices prepared from 14-month-old wild-type mice and memory-impaired APP/PS1 double knock-in mice. Surprisingly, the functional channel density of L-VSCCs was significantly reduced in the APP/PS1 group. No differences in voltage dependency and unitary conductance of L-VSCCs were observed. The results suggest that mechanisms for Ca^2 + dysregulation can differ substantially between animal models of normal aging and models of pathological aging.     

Reduction in neuronal L-type calcium channel activity in a double knock-in mouse model of Alzheimer's disease

Increased function of neuronal L-type voltage-sensitive Ca(2+) channels (L-VSCCs) is strongly linked to impaired memory and altered hippocampal synaptic plasticity in aged rats. However, no studies have directly assessed L-VSCC function in any of the common mouse models of Alzheimer's disease where neurologic deficits are typically more robust. Here, we used cell-attached patch-clamp recording techniques to measure L-VSCC activity in CA1 pyramidal neurons of partially dissociated hippocampal "zipper" slices prepared from 14-month-old wild-type mice and memory-impaired APP/PS1 double knock-in mice. Surprisingly, the functional channel density of L-VSCCs was significantly reduced in the APP/PS1 group. No differences in voltage dependency and unitary conductance of L-VSCCs were observed. The results suggest that mechanisms for Ca(2+) dysregulation can differ substantially between animal models of normal aging and models of pathological aging.     

Beta 2-microglobulin-deficient background ameliorates lethal phenotype of the TGF-beta 1 null mouse.

TGF-beta 1 null (TGF-beta1-/-) mice die at 3-4 wk of age and show an autoimmune inflammatory phenotype associated with enhanced expression of both class I and II MHC molecules. To determine the role of MHC class I Ags in the autoimmune manifestations and the inflammation observed in TGF-beta 1-/- mice, we generated TGF-beta 1-/- mice in the genetic background of beta 2-microglobulin deficiency (beta 2M-/-). TGF-beta 1-/-;beta 2M-/- mice had improved survival compared with TGF-beta 1-/- mice. Histopathological examination showed less severe inflammation, especially in the heart, where Mac-2 reactive macrophages were significantly decreased as compared with TGF-beta 1-/- mice. In vivo depletion of CD8+ T cells in TGF-beta 1-/- mice confirmed suppression of inflammation and reduction in the severity of the wasting syndrome. MHC class II mRNA expression in TGF-beta 1-/-;beta 2M-/- mice was also lower than that in TGF-beta 1-/- mice, suggesting reduced systemic inflammation. Autoimmune response as judged by serum Ab titers to ssDNA and 16/6 Id and by immune complex deposits in kidney was reduced in TGF-beta 1-/-;beta 2M-/- mice, when compared with that in TGF-beta 1-/- mice. Our data thus indicate that MHC class I molecules influence the development of the autoimmunity and the inflammation seen in TGF-beta 1-/- mice and CD8+ T cells may have a contribution to the inflammation in TGF-beta 1-/- mice.     

Genetic modification of the schisis phenotype in a mouse model of X-linked retinoschisis

X-linked retinoschisis (XLRS) is an inherited form of macular degeneration that is caused by mutations in the retinoschisin (RS1) gene. In addition to macular degeneration, other major characteristics of XLRS include splitting of the retina (schisis) and impaired synaptic transmission as indicated by a reduction in the electroretinogram b-wave. It has been known that patients carrying RS1 mutations show a broad range of phenotypic variability. Interestingly, phenotypic variation is observed even among family members with the same RS1 mutation, suggesting the existence of genetic or environmental factors that contribute to the severity of XLRS. However, in the human population, the cause of phenotypic variability and the contribution of genetic modifiers for this relatively rare disease are difficult to study and poorly understood. In this study, using a mouse model for XLRS, we show that genetic factors can contribute to the severity of the retinoschisis phenotype. We report evidence of a major genetic modifier of Rs1, which affects the disease severity in these animals. A quantitative trait locus (QTL), named modifier of Rs1 1 (Mor1), is mapped on chromosome (Chr) 7. When homozygous, the Mor1 allele from the inbred mouse strain AKR/J diminishes the severity of the schisis phenotype in Rs1(tmgc1)/Y male and Rs1(tmgc1)/Rs1(tmgc1) female mice. We also show that the penetrance of the disease phenotype is affected by additional genetic factor(s). Our study suggests that multiple genetic modifiers could potentially be responsible for the phenotypic variation in human XLRS.     

A hypermutator phenotype attenuates the virulence of Listeria monocytogenes in a mouse model

The integrity of the genetic material of bacteria is guaranteed by a set of distinct repair mechanisms. The participation of these repair systems in bacterial pathogenicity has been addressed only recently. Here, we study for the first time the participation in virulence of the MutSL mismatch repair system of Listeria monocytogenes. The mutS and mutL genes, which are contiguous in the L. monocytogenes chromosome, were identified after in silico analysis. The deduced MutS shares 62% identity with MutS of Bacillus subtilis and 50% identity with HexA, its homologue in Streptococcus pneumoniae; MutL shares 59% identity with MutL of B. subtilis and 47% identity with HexB of S. pneumoniae. Functional analysis of the mutSL locus was studied by constructing a double knock-out mutant. We showed that the deletion DeltamutSL induces: (i) a 100- to 1000-fold increase in the spontaneous mutation rate; and (ii) a 10- to 15-fold increase in the frequency of transduction, thus demonstrating the role of mutSL of L. monocytogenes in both mismatch repair and homologous recombination. We found that the deletion DeltamutSL moderately affected bacterial virulence, with a 1-log increase in the lethal dose 50% (LD50) in the mouse. Strikingly, repeated passages of the mutant strain in mice reduced virulence further. Competition assays between wild-type and mutant strains showed that the deletion DeltamutSL reduced the capacity of L. monocytogenes to survive and multiply in mice. These results thus demonstrate that, for the intracellular pathogen L. monocytogenes, a hypermutator phenotype is more deleterious than profitable to its virulence.     

Redox metal chelation ameliorates radiation-induced bone marrow toxicity in a mouse model.

Since zinc desferrioxamine (Zn-DFO) has been shown to be a very potent protector against injuries induced by redox-active metal ions, we examined its protective effect against radiation-induced toxicity. We found that treatment with Zn-DFO given before TBI increased the survival of mice irradiated with 7.5 and 8.5 Gy. Zn-DFO also protected against radiation-induced myelosuppression and body weight loss, while soluble Il6 levels in serum were normalized in mice pretreated with Zn-DFO. We concluded that administration of Zn-DFO prior to TBI protected BALB/c mice from radiation-induced toxicity, increasing survival rates by up to 75%. The biological effect of Zn-DFO is known to result from its effect on the production of intracellular hydroxyl free radicals mediated by redox-active metal ions, and both metal chelation and zinc delivery appear to be equally likely mechanisms for this outcome. We suggest that radiation-induced toxicity is caused by the deleterious effect of redox-active metal ions, and that compounds which modulate this redox activity may act as radioprotectors.     

The tripeptide analog feG ameliorates severity of acute pancreatitis in a caerulein mouse model

Acute pancreatitis (AP) is associated with significant morbidity and mortality; however, there is no specific treatment for this disease. A novel salivary tripeptide analog, feG, reduces inflammation in several different animal models of inflammation. The aims of this study were to determine whether feG reduced the severity of AP and modifies the expression of pancreatic ICAM-1 mRNA during AP in a mouse model. AP was induced in mice by hourly (x12) intraperitoneal injections of caerulein. A single dose of feG (100 microg/kg) was coadministered with caerulein either at time 0 h (prophylactic) or 3 h after AP induction (therapeutic). Plasma amylase and pancreatic MPO activities and pancreatic ICAM-1 mRNA expression (by RT-PCR) were measured. Pancreatic sections were histologically assessed for abnormal acinar cells and interstitial space. AP induction produced a sevenfold increase in plasma amylase, a tenfold increase in pancreatic MPO activity, and a threefold increase in interstitial space, and 90% of the acinar cells were abnormal. Prophylactic treatment with feG reduced the AP-induced plasma amylase activity by 45%, pancreatic MPO by 80%, the proportion of abnormal acinar cells by 30%, and interstitial space by 40%. Therapeutic treatment with feG significantly reduced the AP-induced abnormal acinar cells by 10% and the interstitial space by 20%. Pancreatic ICAM-1 mRNA expression was upregulated in AP and was reduced by 50% with prophylactic and therapeutic treatment with feG. We conclude that feG ameliorates experimental AP acting at least in part by modulating ICAM-1 expression in the pancreas.     

Oridonin ameliorates neuropathological changes and behavioural deficits in a mouse model of cerebral amyloidosis

Alzheimer's disease (AD) is the most common form of neurodegeneration and the major cause of dementia. This multifactorial disorder is clinically defined by progressive behavioural and cognitive deficits, and neuropathologically characterized by β‐amyloid aggregation, hyperphosphorylated tau and neuroinflammation. Oridonin, a diterpenoid isolated from Chinese herb Rabdosia rubescens, has multiple biological properties, especially anti‐inflammatory and neuroregulatory activities. Potential therapeutic effects of Oridonin were investigated in an animal model of cerebral amyloidosis for AD, transgenic APP/PS1 mice. Oridonin was suspended in carboxymethylcellulose or loaded with a nanostructured emulsion, and was orally administrated or injected. Before, during and following the experimental treatments, behavioural tests were performed with these transgenic mice and their naive littermates. Following relatively short‐term treatments of 10 days, brain tissue of mice were removed for immunohistochemical assays. The results indicate that both oral treatment and injection of Oridonin significantly attenuated β‐amyloid deposition, plaque‐associated APP expression and microglial activation in brain of transgenic mice. Furthermore, injection of Oridonin‐nanoemulsion ameliorated deficits in nesting, an important affiliative behaviour, and in social interaction. Additional in vitro studies indicated that Oridonin effectively attenuated inflammatory reaction of macrophage and microglial cell lines. Our results suggest that Oridonin might be considered a promising therapeutic option for human AD or other neurodegenerative diseases.     

Williams-Beuren syndrome: a model of recurrent genomic mutation

Williams-Beuren syndrome is a segmental aneusomy syndrome with manifestations affecting the vascular, connective tissue, endocrine and central nervous systems. Most patients show a similar heterozygous approximately 1.5 Mb deletion at 7q11.23 that contains a number of reported genes. Deletion mapping in the few atypical patients with smaller deletions suggested that additive effects of haploinsufficiency for two or more genes might be necessary for the phenotype. Vascular stenoses are caused by haploinsufficiency at the elastin gene, while the genes responsible for the cognitive deficits are likely located at the telomeric edge of the deletion, including CYLN2 and GTF2I. Large region-specific segmental duplications predispose to misalignment and inter- or intrachromosomal unequal crossing-over causing the deletions. Atypical alleles at 7q11.23 such as inversions and deletions/insertions of large repeats, also generated through aberrant recombination between the local segmental duplications, are found in approximately 35% of transmitting parents. Genomic instability at 7q11.23 is directly related to the genomic structure of the region.     

Genetic background modulates the phenotype of a mouse model of DYT1 dystonia

DYT1 dystonia is a debilitating neurological disease characterized by involuntary twisting movements. The disease is caused by an in-frame deletion (GAG, "ΔE") mutation in the TOR1A gene that encodes the torsinA protein. Intriguingly, only 30% of mutation carriers exhibit motor symptoms despite the fact that functional brain imaging studies show abnormal brain metabolism in all carriers. Because genetic modifiers may be a determinant of this reduced penetrance, we examined the genetic contribution of three different inbred strains of mice on the DYT1 mutation in animals that are homozygous (Tor1a(ΔE/ΔE)) or heterozygous (Tor1a(ΔE/+); disease state) for the disease-causing ΔE mutation. We find that the DBA/2J, C57BL/6J, and CD1-ICR contribution of genes significantly alter lifespan in Tor1a(ΔE/ΔE) mice, which die during the first few days of life on the 129S6/SvEvTac (129) background. The C57BL/6J (B6) strain significantly decreases life expectancy of Tor1a(ΔE/ΔE) animals but, like 129S6/SvEvTac Tor1a(ΔE/+) mice, congenic C57BL/6J Tor1a(ΔE/+) mice do not exhibit any motor abnormalities. In contrast, the DBA/2J (D2) strain significantly increases life expectancy. This effect was not present in congenic DBA/2J Tor1a(ΔE/ΔE) mice, indicating that the extended lifespan of F2 129/D2 mice was due to a combination of homozygous and heterozygous allelic effects. Our observations suggest that genetic modifiers may alter the penetrance of the ΔE mutation, and that mapping these modifiers may provide fresh insight into the torsinA molecular pathway.     

Ascorbic acid treatment corrects the phenotype of a mouse model of Charcot-Marie-Tooth disease

Charcot-Marie-Tooth disease (CMT) is the most common hereditary peripheral neuropathy, affecting 1 in 2,500 people. The only treatment currently available is rehabilitation or corrective surgery. The most frequent form of the disease, CMT-1A, involves abnormal myelination of the peripheral nerves. Here we used a mouse model of CMT-1A to test the ability of ascorbic acid, a known promoter of myelination, to correct the CMT-1A phenotype. Ascorbic acid treatment resulted in substantial amelioration of the CMT-1A phenotype, and reduced the expression of PMP22 to a level below what is necessary to induce the disease phenotype. As ascorbic acid has already been approved by the FDA for other clinical indications, it offers an immediate therapeutic possibility for patients with the disease.     

Hepatocyte growth factor ameliorates dermal sclerosis in the tight-skin mouse model of scleroderma

The tight-skin (TSK/+) mouse, a genetic model of systemic sclerosis (SSc), develops cutaneous fibrosis and defects in pulmonary architecture. Because hepatocyte growth factor (HGF) is an important mitogen and morphogen that contributes to the repair process after tissue injury, we investigated the role of HGF in cutaneous fibrosis and pulmonary architecture defects in SSc using TSK/+ mice. TSK/+ mice were injected in the gluteal muscle with either hemagglutinating virus of Japan (HVJ) liposomes containing 8 μg of a human HGF expression vector (HGF-HVJ liposomes) or a mock vector (untreated control). Gene transfer was repeated once weekly for 8 weeks. The effects of HGF gene transfection on the histopathology and expression of tumor growth factor (TGF)-β and IL-4 mRNA in TSK/+ mice were examined. The effect of recombinant HGF on IL-4 production by TSK/+ CD4⁺ T cells stimulated by allogeneic dendritic cells (DCs) in vitro was also examined. Histologic analysis revealed that HGF gene transfection in TSK/+ mice resulted in a marked reduction of hypodermal thickness, including the subcutaneous connective tissue layer. The hypodermal thickness of HGF-treated TSK/+ mice was decreased two-fold to three-fold compared with untreated TSK/+ mice. However, TSK/+ associated defects in pulmonary architecture were unaffected by HGF gene transfection. HGF gene transfection significantly inhibited the expression of IL-4 and TGF-β1 mRNA in the spleen and skin but not in the lung. We also performed a mixed lymphocyte culture and examined the effect of recombinant HGF on the generation of IL-4. Recombinant HGF significantly inhibited IL-4 production in TSK/+ CD4⁺ T cells stimulated by allogeneic DCs. HGF gene transfection inhibited IL-4 and TGF-β mRNA expression, which has been postulated to have a major role in fibrinogenesis and reduced hypodermal thickness, including the subcutaneous connective tissue layer of TSK/+ mice. HGF might represent a novel strategy for the treatment of SSc.     

Hepatocyte growth factor ameliorates dermal sclerosis in the tight-skin mouse model of scleroderma

The tight-skin (TSK/+) mouse, a genetic model of systemic sclerosis (SSc), develops cutaneous fibrosis and defects in pulmonary architecture. Because hepatocyte growth factor (HGF) is an important mitogen and morphogen that contributes to the repair process after tissue injury, we investigated the role of HGF in cutaneous fibrosis and pulmonary architecture defects in SSc using TSK/+ mice. TSK/+ mice were injected in the gluteal muscle with either hemagglutinating virus of Japan (HVJ) liposomes containing 8 mug of a human HGF expression vector (HGF-HVJ liposomes) or a mock vector (untreated control). Gene transfer was repeated once weekly for 8 weeks. The effects of HGF gene transfection on the histopathology and expression of tumor growth factor (TGF)-beta and IL-4 mRNA in TSK/+ mice were examined. The effect of recombinant HGF on IL-4 production by TSK/+ CD4+ T cells stimulated by allogeneic dendritic cells (DCs) in vitro was also examined. Histologic analysis revealed that HGF gene transfection in TSK/+ mice resulted in a marked reduction of hypodermal thickness, including the subcutaneous connective tissue layer. The hypodermal thickness of HGF-treated TSK/+ mice was decreased two-fold to three-fold compared with untreated TSK/+ mice. However, TSK/+ associated defects in pulmonary architecture were unaffected by HGF gene transfection. HGF gene transfection significantly inhibited the expression of IL-4 and TGF-beta1 mRNA in the spleen and skin but not in the lung. We also performed a mixed lymphocyte culture and examined the effect of recombinant HGF on the generation of IL-4. Recombinant HGF significantly inhibited IL-4 production in TSK/+ CD4+ T cells stimulated by allogeneic DCs. HGF gene transfection inhibited IL-4 and TGF-beta mRNA expression, which has been postulated to have a major role in fibrinogenesis and reduced hypodermal thickness, including the subcutaneous connective tissue layer of TSK/+ mice. HGF might represent a novel strategy for the treatment of SSc.