Loss-of-function variants in myocardin cause congenital megabladder in humans and mice

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Abstract

Myocardin (MYOCD) is the founding member of a class of transcriptional co-activators that bind serum response factor to activate gene expression programs critical in smooth muscle (SM) and cardiac muscle development. Insights into the molecular functions of MYOCD have been obtained from cell culture studies and, to date, knowledge about in vivo roles of MYOCD comes exclusively from experimental animals. Here, we defined an often lethal congenital human disease associated with inheritance of pathogenic MYOCD variants. This disease manifested as a massively dilated urinary bladder, or megabladder, with disrupted SM in its wall. We provided evidence that monoallelic loss-of-function variants in MYOCD caused congenital megabladder in males only, whereas biallelic variants were associated with disease in both sexes, with a phenotype additionally involving the cardiovascular system. These results were supported by co-segregation of MYOCD variants with the phenotype in four unrelated families, by in vitro transactivation studies where pathogenic variants resulted in abrogated SM gene expression, and finding megabladder in two distinct mouse models with reduced Myocd activity. In conclusion, we have demonstrated that variants in MYOCD result in human disease, and the collective findings highlight a vital role for MYOCD in mammalian organogenesis.

Authors

Arjan C. Houweling, Glenda M. Beaman, Alex V. Postma, T. Blair Gainous, Klaske D. Lichtenbelt, Francesco Brancati, Filipa M. Lopes, Ingeborg van der Made, Abeltje M. Polstra, Michael L. Robinson, Kevin D. Wright, Jamie M. Ellingford, Ashley R. Jackson, Eline Overwater, Rita Genesio, Silvio Romano, Letizia Camerota, Emanuela D'Angelo, Elizabeth J. Meijers-Heijboer, Vincent M. Christoffels, Kirk M. McHugh, Brian L. Black, William G. Newman, Adrian S. Woolf, Esther E. Creemers

Dermal adipose tissue has high plasticity and undergoes reversible dedifferentiation in mice

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Abstract

Dermal adipose tissue (dWAT) has been the focus of much discussion in recent years. However, dWAT remains poorly characterized. The fate of the mature dermal adipocytes and the origin of the rapidly re-appearing dermal adipocytes at different stages remain unclear. Here, we isolated dermal adipocytes and characterized dermal fat at the cellular and molecular level. Together with its dynamic responses to external stimuli, we established that dermal adipocytes are a distinct class of white adipocytes with high plasticity. By combining pulse-chase lineage tracing and single cell RNA-sequencing, we observed that mature dermal adipocytes undergo de-differentiation and re-differentiation under physiological and pathophysiological conditions. Upon various challenges, the de-differentiated cells proliferate and re-differentiate into adipocytes. In addition, manipulation of dWAT highlighted an important role for mature dermal adipocytes for hair cycling and wound healing. Altogether, these observations unravel a surprising plasticity of dermal adipocytes and provide an explanation for the dynamic changes in dWAT mass that occur under physiological and pathophysiological conditions, and highlight the important contributions of dWAT towards maintaining skin homeostasis.

Authors

Zhuzhen Zhang, Mengle Shao, Chelsea Hepler, Zhenzhen Zi, Shangang Zhao, Yu A. An, Yi Zhu, Alexandra Ghaben, May-yun Wang, Na Li, Toshiharu Onodera, Nolwenn Joffin, Clair Crewe, Qingzhang Zhu, Lavanya Vishvanath, Ashwani Kumar, Chao Xing, Qiong A. Wang, Laurent Gautron, Yingfeng Deng, Ruth Gordillo, Ilja Kruglikov, Christine M. Kusminski, Rana K. Gupta, Philipp E. Scherer

Cannabidiol attenuates seizures and EEG abnormalities in Angelman syndrome model mice

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Abstract

Angelman syndrome (AS) is a neurodevelopmental disorder characterized by intellectual disability, lack of speech, ataxia, EEG abnormalities, and epilepsy. Seizures in AS individuals are common, debilitating, and often drug-resistant. Therefore, there is an unmet need for better treatment options. Cannabidiol (CBD), a major phytocannabinoid constituent of cannabis, has antiseizure activity and behavioral benefits in preclinical and clinical studies for some disorders associated with epilepsy, suggesting that the same could be true for AS. Here we show that acute CBD (100 mg/kg) attenuated hyperthermia- and acoustically-induced seizures in a mouse model of AS. However, neither acute CBD nor a two-weeklong course of CBD administered immediately after a kindling protocol could halt the pro-epileptogenic plasticity observed in AS model mice. CBD had a dose-dependent sedative effect, but did not have an impact on motor performance. CBD abrogated the enhanced intracortical local field potential power, including delta and theta rhythms observed in AS model mice, indicating that CBD administration could also help normalize the EEG deficits observed in individuals with AS. Our results provide critical preclinical evidence supporting CBD treatment of seizures and alleviation of EEG abnormalities in AS, and will thus help guide the rational development of CBD as an AS treatment.

Authors

Bin Gu, Manhua Zhu, Madison R. Glass, Marie Rougié, Viktoriya D. Nikolova, Sheryl S. Moy, Paul R. Carney, Benjamin D. Philpot

Myeloid loss of Beclin 1 promotes PD-L1^hi precursor B Cell lymphoma development

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Abstract

Beclin 1 (Becn1) is a key molecule of the autophagy pathway and has been implicated in cancer development. Due to the embryonic lethality of Becn1 homozygous deficient mice, the precise mechanisms and cell-type-specific role of Becn1 in the regulation of inflammation and tumor immunity remain elusive. Here, we report that myeloid-deficient Becn1 (Becn1ΔM) mice develop neutrophilia and hypersusceptible to LPS-induced septic shock, with a high risk of developing spontaneous precursor (pre)-B cell lymphoma with elevated expressions of immunosuppressive molecules PD-L1 and IL-10. Becn1 deficiency results in stabilization of neutrophil MEKK3, aberrant p38 activation, and neutrophil-B cell interaction through Cxcl9/Cxcr3 chemotaxis. Neutrophil-B cell interplay leads to activations of IL-21/STAT3/IRF1 and CD40L/ERK signaling, together regulates the programmed death ligand 1 (PD-L1) expression, and suppresses CD8+ T cell function. Ablation of p38 in Becn1ΔM mice prevents neutrophil-inflammation and B cell tumorigenesis. Importantly, low Becn1 expression in human neutrophils correlates with PD-L1 levels in pre-B ALL patients. Our findings have identified myeloid Becn1 as a therapeutic target of cancer immunity and immunotherapy for pre-B lymphomas.

Authors

Peng Tan, Lian He, Changsheng Xing, Jingrong Mao, Xiao Yu, Motao Zhu, Lixia Diao, Leng Han, Yubin Zhou, James M. You, Helen Y. Wang, Rong-Fu Wang

Sonic Hedgehog repression underlies gigaxonin mutation-induced motor deficits in giant axonal neuropathy

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Abstract

Growing evidence shows that alterations occurring at early developmental stages contribute to symptoms manifested in adulthood in the setting of neurodegenerative diseases. Here, we studied the molecular mechanisms causing giant axonal neuropathy (GAN), a severe neurodegenerative disease due to loss-of-function of the gigaxonin-E3 ligase. We showed that gigaxonin governs Sonic Hedgehog (Shh) induction, the developmental pathway patterning the dorso-ventral axis of the neural tube and muscles, by controlling the degradation of the Shh-bound Patched receptor. Similarly to Shh inhibition, repression of gigaxonin in zebrafish impaired motor neuron specification and somitogenesis and abolished neuromuscular junction formation and locomotion. Shh signaling was impaired in gigaxonin null zebrafish and was corrected by both pharmacological activation of the Shh pathway and human gigaxonin, pointing to an evolutionary-conserved mechanism regulating Shh signaling. Gigaxonin-dependent inhibition of Shh activation was also demonstrated in primary fibroblasts from GAN patients and in a Shh activity reporter line depleted in gigaxonin. Our findings establish gigaxonin as a key E3 ligase that positively controls the initiation of Shh transduction, reveal the causal role of Shh dysfunction in motor deficits, thus highlighting the developmental origin of GAN.

Authors

Yoan Arribat, Karolina S Mysiak, Léa Lescouzères, Alexia Boizot, Maxime Ruiz, Mireille Rossel, Pascale Bomont

Expression of mitochondrial membrane-linked SAB determines severity of sex-dependent acute liver injury

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Abstract

SAB is an outer membrane docking protein for JNK mediated impaired mitochondrial function. Deletion of Sab in hepatocytes inhibits sustained JNK activation and cell death. Current work demonstrated that increasing SAB enhanced the severity of APAP liver injury. Female mice were resistant to liver injury and exhibited markedly decreased hepatic SAB protein expression versus males. The mechanism of SAB repression involved a pathway from ERα to p53 expression which induced miR34a-5p. miR34a-5p targeted the Sab mRNA coding region, repressing SAB expression. Fulvestrant or p53 knockdown decreased miR34a-5p and increased SAB in females leading to increased injury from APAP and TNF/galactosamine. In contrast, ERα agonist increased p53 and miR34a-5p which decreased SAB expression and hepatotoxicity in males. Hepatocyte-specific deletion of miR34a also increased severity of liver injury in females, which was prevented by GalNAc-ASO knockdown of Sab. Similar to mice, premenopausal human females also expressed high hepatic p53 and low SAB levels while age-matched males expressed low p53 and high SAB levels, but there was no sex difference of SAB expression in postmenopause. In conclusion, the level of SAB expression determined the severity of JNK dependent liver injury. Females expressed low hepatic SAB protein levels due to an ERα-p53-miR34a pathway which repressed SAB expression, accounting for resistance to liver injury.

Authors

Sanda Win, Robert W.M. Min, Christopher Q. Chen, Jun Zhang, Yibu Chen, Meng Li, Ayako Suzuki, Manal F. Abdelmalek, Ying Wang, Mariam Aghajan, Filbert W.M. Aung, Anna Mae Diehl, Roger J. Davis, Tin A. Than, Neil Kaplowitz

Schwann cells expressing nociceptive channel TRPA1 orchestrate ethanol-evoked neuropathic pain in mice

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Abstract

Excessive alcohol consumption is associated with spontaneous burning pain, hyperalgesia and allodynia. Although acetaldehyde has been implicated in the painful alcoholic neuropathy, the mechanism by which the ethanol metabolite causes pain symptoms is unknown. Acute ethanol ingestion caused delayed mechanical allodynia in mice. Inhibition of alcohol dehydrogenase (ADH) or deletion of transient receptor potential ankyrin 1 (TRPA1), a sensor for oxidative and carbonyl stress, prevented allodynia. Acetaldehyde generated by ADH in both liver and Schwann cells surrounding nociceptors was required for TRPA1-induced mechanical allodynia. Plp1-Cre;Trpa1fl/fl mice with a tamoxifen-inducible specific deletion of TRPA1 in Schwann cells revealed that channel activation by acetaldehyde in these cells initiates a NADPH oxidase-1 (NOX-1)-dependent production of hydrogen peroxide (H2O2) and 4-hydroxynonenal (4-HNE), which sustains allodynia by paracrine targeting of nociceptor TRPA1. Chronic ethanol ingestion caused prolonged mechanical allodynia and loss of intraepidermal small nerve fibers in WT mice. While Trpa1-/- or Plp1-Cre;Trpa1fl/fl mice did not develop mechanical allodynia, they did not show any protection from the small fiber neuropathy. Human Schwann cells express ADH/TRPA1/NOX1 and recapitulate the proalgesic functions of mouse Schwann cells. TRPA1 antagonists might attenuate some symptoms of alcohol-related pain.

Authors

Francesco De Logu, Simone Li Puma, Lorenzo Landini, Francesca Portelli, Alessandro Innocenti, Daniel Souza Monteiro de Araújo, Malvin N. Janal, Riccardo Patacchini, Nigel W. Bunnett, Pierangelo Geppetti, Romina Nassini

CCL28-induced RARβ expression inhibits oral squamous cell carcinoma bone invasion

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Abstract

Oral squamous cell carcinoma (OSCC) frequently invades the maxillary or mandibular bone, and this bone invasion is closely associated with poor prognosis and survival. Here, we show that CCL28 functions as a negative regulator of OSCC bone invasion. CCL28 inhibited invasion and epithelial-mesenchymal transition (EMT), and its inhibition of EMT was characterized by induced E-cadherin expression and reduced nuclear localization of beta-catenin in OSCC cells with detectable RUNX3 expression levels. CCL28 signaling via CCR10 increased retinoic acid receptor (RAR)β expression by reducing the interaction between RARα and HDAC1. In addition, CCL28 reduced RANKL production in OSCC and osteoblastic cells and blocked RANKL-induced osteoclastogenesis in osteoclast precursors. Intraperitoneally administered CCL28 inhibited tumor growth and osteolysis in mouse calvaria and tibia inoculated with OSCC cells. RARβ expression was also increased in tumor tissues. In OSCC patients, low CCL28, CCR10, and RARβ expression levels were highly correlated with bone invasion. OSCC patients with higher expression of CCL28, CCR10, or RARβ had significantly better overall survival. These findings suggest that CCL28, CCR10, and RARβ are useful markers for the prediction and treatment of OSCC bone invasion. Furthermore, CCL28 upregulation in OSCC cells or CCL28 treatment can be a therapeutic strategy for OSCC bone invasion.

Authors

Junhee Park, Xianglan Zhang, Sun Kyoung Lee, Na-Young Song, Seung Hwa Son, Ki Rim Kim, Jae Hoon Shim, Kwang-Kyun Park, Won-Yoon Chung

Splicing factor SRSF1 controls T cell hyperactivity and systemic autoimmunity

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Abstract

Systemic lupus erythematosus (SLE) is a devastating autoimmune disease, in which hyperactive T cells play a critical role. Understanding molecular mechanisms underlying the T cell hyperactivity will lead to identification of specific therapeutic targets. Serine/arginine-rich splicing factor (SRSF)1 is an essential RNA-binding protein which controls posttranscriptional gene expression. We have demonstrated that SRSF1 levels are aberrantly decreased in T cells from SLE patients and correlate with severe disease, yet the role of SRSF1 in T cell physiology and autoimmune disease is largely unknown. Here we show that T cell-restricted Srsf1-deficient mice develop systemic autoimmunity and lupus-nephritis. Mice exhibit increased frequencies of activated/effector T cells producing proinflammatory cytokines, and an elevated T cell activation gene signature. Mechanistically, we noted increased activity of the mechanistic target of rapamycin (mTOR) pathway and reduced expression of its repressor PTEN. The mTOR complex (mTORC)1 inhibitor rapamycin suppressed proinflammatory cytokine production by T cells and alleviated autoimmunity in Srsf1-deficient mice. Of direct clinical relevance, PTEN levels correlated with SRSF1 in T cells from SLE patients, and SRSF1 overexpression rescued PTEN, suppressed mTORC1 activation and proinflammatory cytokine production. Our studies reveal the role of a previously unrecognized molecule SRSF1 in restraining T cell activation and averting the development of autoimmune disease and a potential therapeutic target for lupus.

Authors

Takayuki Katsuyama, Hao Li, Denis Comte, George C. Tsokos, Vaishali R. Moulton

Soluble epoxide hydrolase promotes astrocyte survival in retinopathy of prematurity

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Abstract

Polyunsaturated fatty acids (PUFAs) such as docosahexaenoic acid (DHA) positively affect the outcome of retinopathy of prematurity (ROP). Given that DHA metabolism by cytochrome P450 and soluble epoxide hydrolase (sEH) enzymes affects retinal angiogenesis and vascular stability we investigated the role of sEH in a mouse model of ROP. In wild-type mice, hyperoxia elicited the tyrosine nitration and inhibition of the sEH and decreased generation of the DHA-derived diol 19,20-dihydroxydocosapentaenoic acid (DHDP). Correspondingly in a murine model of ROP, sEH–/– mice developed a larger central avascular zone and peripheral pathological vascular tuft formation than their wild-type littermates. Astrocytes were the cells most affected by sEH deletion and hyperoxia increased astrocyte apoptosis. In rescue experiments 19,20-DHDP prevented astrocyte loss by targeting the mitochondrial membrane to prevent the hyperoxia-induced dissociation of presenilin-1 (PS-1) and PS-1 associated protein (PSAP) to attenuate PARP1 activation and mitochondrial DNA damage. Therapeutic intravitreal administration of 19,20-DHDP not only suppressed astrocyte loss but also reduced pathological vascular tuft formation in sEH–/– mice. Our data indicate that sEH activity is required for mitochondrial integrity and retinal astrocyte survival in ROP. Moreover, 19,20-DHDP may be more effective than DHA as a nutritional supplement at preventing retinopathy in preterm infants.

Authors

Jiong Hu, Sofia Iris Bibli, Janina Wittig, Sven Zukunft, Jihong Lin, Hans-Peter Hammes, Rüdiger Popp, Ingrid Fleming