An infection model for SARS-CoV-2 using rat transplanted with hiPSC-airway epithelial cells.

Investigating the infection mechanism of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) in the airway epithelium and developing effective defense strategies against infection are important. To achieve this, establishing appropriate infection models is crucial. Therefore various in vitro models, such as cell lines and primary cultures, and in vivo models involving animals that exhibit SARS-CoV-2 infection and genetically humanized animals, have been used as animal models. However, no animal model has been established that allows infection experiments with human cells under the physiological environment of airway epithelia. Therefore, we aimed to establish a novel animal model that enables infection experiments using human cells. Human iPSC-derived airway epithelial cell-transplanted nude rats (hiPSC-AEC rats) were used, and infection studies were performed by spraying lentiviral pseudoviruses containing SARS-CoV-2 spike protein and the GFP gene on the tracheae. After infection, immunohistochemical analyses revealed the existence of GFP-positive infected transplanted cells in the epithelial and submucosal layers. In this study, a SARS-CoV-2 infection animal model including human cells was established mimicking infection through respiration and we demonstrated that the hiPSC-AEC rat could be used as an animal model for basic research and the development of therapeutic methods for human-specific respiratory infectious diseases.

Fatal Community-Acquired Clostridioides difficile Infection as a Cause of Listeria Meningitis: A Case Report.

A 77-year-old woman with a history of total gastrectomy was transferred to our hospital with complaints of fever and consciousness disturbance for five days. She had fever and consciousness disturbance with positive meningeal signs. Laboratory findings indicated an elevated inflammatory response and hypoalbuminemia, and computed tomography (CT) of the body indicated intestinal gas retention and mild ascites. Cerebrospinal fluid analysis revealed pleocytosis with elevated protein levels and a diagnosis of Listeria meningitis was made. Treatment with ampicillin/sulbactam was started, and her fever and consciousness disturbance resolved on day 2. However, on day 3, her fever and conscious disturbance deteriorated, and she went into shock subsequently. Laboratory findings revealed deteriorated inflammatory response and hypoalbuminemia. Body CT showed an obvious distended bowel loop and intestinal edema. A stool culture revealed positive Clostridioides difficile toxin B, and we diagnosed her with Clostridioides difficile infection (CDI). Although intravenous metronidazole was initiated, she died due to prolonged hypovolemic shock. We considered she had community-acquired CDI because her CDI emerged immediately after the initiation of antibiotics, symptom deterioration within 48 hours of admission, and abnormal abdominal CT findings at admission. Listeria meningitis can develop based on community-acquired CDI. Because CDI can have a very rapid and fatal course and is sometimes complicated by other infectious diseases, clinicians should pay attention to this complication.

In vivo CRISPR screening directly targeting testicular cells.

CRISPR-Cas9 short guide RNA (sgRNA) library screening is a powerful approach to understand the molecular mechanisms of biological phenomena. However, its in vivo application is currently limited. Here, we developed our previously established in vitro revival screening method into an in vivo one to identify factors involved in spermatogenesis integrity by utilizing sperm capacitation as an indicator. By introducing an sgRNA library into testicular cells, we successfully pinpointed the retinal degeneration 3 (Rd3) gene as a significant factor in spermatogenesis. Single-cell RNA sequencing (scRNA-seq) analysis highlighted the high expression of Rd3 in round spermatids, and proteomics analysis indicated that Rd3 interacts with mitochondria. To search for cell-type-specific signaling pathways based on scRNA-seq and proteomics analyses, we developed a computational tool, Hub-Explorer. Through this, we discovered that Rd3 modulates oxidative stress by regulating mitochondrial distribution upon ciliogenesis induction. Collectively, our screening system provides a valuable in vivo approach to decipher molecular mechanisms in biological processes.

A case of meningoencephalitis caused by multisystem inflammatory syndrome in adult SARS-CoV-2 infection.

A 37-year-old woman was hospitalized with fever and consciousness disturbance. She showed systemic inflammation with stress cardiomyopathy. Brain computed tomography showed diffuse brain edema. Cerebrospinal fluid (CSF) findings revealed markedly elevated cerebrospinal fluid pressure with pleocytosis, elevated protein, and elevated interleukin 6. The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nicking enzyme amplification reaction test using a nasopharyngeal swab was positive, and the patient was diagnosed with SARS-CoV-2 infection. From the negative result of the CSF SARS-CoV-2 polymerase chain reaction test and no findings of bacterial or viral infection, we diagnosed meningoencephalitis by multisystem inflammation syndrome in adults (MIS-A). Intravenous methylprednisolone pulse therapy improved her symptoms and brain edema. There have been no cases of MIS-A with meningoencephalitis, and no initial treatment strategy has been established, especially in emergency cases of suspected MIS-A. The present case suggested Early intravenous methylprednisolone pulse with anti-coronaviral therapies after the exclusion of bacterial infection would be useful in suspected MIS-A with emergent meningoencephalitis cases.

Enteral tube nutrition for geriatric post-stroke dysphagia evaluation (ENGE) score to evaluate the risk of dysphagia after acute ischemic stroke.

BACKGROUND: Post-stroke dysphagia (PSD) is a common complication after stroke. Early PSD prediction is essential for patient stratification for intensive oral intake rehabilitation. We aimed to develop a PSD prediction score using clinical data obtained at admission. METHODS: We examined consecutive patients with acute ischemic stroke between 2018 and 2019. The dysphagia status 14 days after admission was assessed using the Functional Oral Intake Scale (FOIS). PSD was defined as FOIS 1-3, which represents tube-dependent nutrition. Using multivariable logistic regression analysis, we constructed the Enteral tube Nutrition for Geriatric post-stroke dysphagia Evaluation (ENGE) score. The discriminative performance of the ENGE score was analyzed by receiver operating curve analysis. The reproducibility of the ENGE score was validated using patient data in 2020. RESULTS: PSD developed in 84 of 488 patients (median age 78 years; 57% males). The ENGE score ranged from 0 to 6, with 1 point assigned for older age (≥78 years), 1 for high premorbid modified Rankin Scale (mRS) (≥1), 3 for high NIHSS score (≥12), and 1 for low serum albumin (<3.0 mg/dl). The area under the curve (AUC) of the ENGE score for discriminating PSD was 0.88 (95% confidence interval [CI] 0.83-0.92), and a score of 3 or more had a higher positive likelihood ratio. In the validation cohort, the AUC of the ENGE score for PSD was 0.85 (95% CI 0.78-0.91), which was similar to the derivation cohort (p = 0.491). CONCLUSIONS: The ENGE score predicts severe PSD after acute ischemic stroke with good reproducibility.

Eustachian Valve-Enhanced Paradoxical Cerebral Embolism: A Case Report.

Cryptogenic stroke includes many suspicious embolic causes that do not fulfill the Trial of Org 10172 in Acute Stroke Treatment (TOAST) classification criteria. Atrial fibrillation (AF) is one of the major hidden causes of cryptogenic stroke, and an implantable loop recorder (ILR) is widely used for detecting AF. Herein, we report a case of paradoxical cerebral embolism due to a large Eustachian valve with large PFO under no molecular complete remission (CR) of acute monocytic leukemia (AMoL). A 75-year-old man arrived at our emergency room because of aphasia and right hemiparesis. He had a history of two cryptogenic strokes and implanted ILR. Magnetic resonance imaging showed left middle cerebral artery occlusion with slight acute ischemic lesion. The red clot was retrieved by mechanical thrombectomy, and complete recanalization was achieved. We checked ILR, but there was no AF. Transesophageal echocardiography revealed a large patent foramen ovale (PFO) and the large Eustachian valve in the right atrium. Although obvious deep vein thrombosis (DVT) was not detected in venous ultrasonography of the lower extremities, Wilms' tumor 1 messenger ribonucleic acid (WT1mRNA) expression level was high, and AMoL was considered to be not in molecular CR, suggesting a high risk of thrombosis to the large Eustachian valve. From large PFO and no molecular CR of AMoL, we diagnosed him with paradoxical cerebral embolism. Ruling out of AF by ILR and other etiologies, such as aortic or carotid atherosclerosis and pulmonary shunt, also supported the diagnosis of paradoxical cerebral embolism. Even in the absence of obvious DVT, paradoxical cerebral embolism should be considered in cases of a large Eustachian valve and PFO with a hypercoagulable state.

Geriatric nutrition risk index predicts prolonged post-stroke dysphagia in acute ischemic stroke.

BACKGROUND: Post-stroke dysphagia (PSD) is a common complication after stroke. Malnutrition inhibits stroke recovery and is associated with stroke mortality. However, no studies have investigated the effects of nutritional state at admission on prolonged PSD. METHODS: We retrospectively analyzed ischemic stroke patients in our institute from January 2018 to December 2020. Swallowing function was assessed using the Food Oral Intake Scale; prolonged PSD was defined as levels 1-3 at 14 days after admission. The Geriatric Nutritional Risk Index (GNRI) was used to assess nutritional risks, which were classified as follows: >98, no nutritional risk; 92-98, mild nutritional risk; 82-92, moderate nutritional risk; and <82, severe nutritional risk. The association between GNRI and prolonged PSD was assessed. RESULTS: Of 580 patients (median age, 81 years; male, 53%), prolonged PSD was detected in 117 patients. Patients with severe dysphagia had older age, higher pre-stroke modified Rankin Scale score, lower GNRI, and higher National Institutes of Health Stroke Scale score. Logistic regression analysis revealed that lower GNRI was independently associated with prolonged PSD (continuous value; adjusted odds ratio [OR] 1.03, 95% confidence interval [CI] 1.00-1.05). In addition, when "severe" and "moderate" nutritional risk was analyzed as a single class, moderate or severe nutritional risk (GNRI < 92) was independently associated with prolonged PSD (adjusted OR 2.50, 95% CI 1.29-4.87), compared with no nutritional risk patients (GNRI > 98). CONCLUSIONS: In acute ischemic stroke, lower GNRI at admission was independently associated with prolonged PSD, suggesting that GNRI at admission might identify patients at risk of prolonged PSD.

CGG repeat expansion in LRP12 in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by the degeneration of motor neurons. Although repeat expansion in C9orf72 is its most common cause, the pathogenesis of ALS isn't fully clear. In this study, we show that repeat expansion in LRP12, a causative variant of oculopharyngodistal myopathy type 1 (OPDM1), is a cause of ALS. We identify CGG repeat expansion in LRP12 in five families and two simplex individuals. These ALS individuals (LRP12-ALS) have 61-100 repeats, which contrasts with most OPDM individuals with repeat expansion in LRP12 (LRP12-OPDM), who have 100-200 repeats. Phosphorylated TDP-43 is present in the cytoplasm of iPS cell-derived motor neurons (iPSMNs) in LRP12-ALS, a finding that reproduces the pathological hallmark of ALS. RNA foci are more prominent in muscle and iPSMNs in LRP12-ALS than in LRP12-OPDM. Muscleblind-like 1 aggregates are observed only in OPDM muscle. In conclusion, CGG repeat expansions in LRP12 cause ALS and OPDM, depending on the length of the repeat. Our findings provide insight into the repeat length-dependent switching of phenotypes.

A Case Report of Deep Cerebral Venous Thrombosis Presenting Unilateral Lesion: The Association of Asymmetric Venous Outflow and Unilateral Lesion.

A 60-year-old man was admitted to our hospital due to progressive aphasia and right hemiparesis. Brain magnetic resonance imaging showed the left thalamus and basal ganglia lesion. Digital subtraction angiography showed the vein of Galen and straight sinus occlusion, suggesting cerebral venous thrombosis. Since his left transverse sinus was hypoplastic, his left deep cerebral lesion was due to the left deep cerebral vein congestion by the asymmetrical venous outflow. After anticoagulant therapy, his symptom and unilateral lesion improved. Clinicians should consider the vein of Galen and straight sinus thrombosis even in unilateral deep cerebral lesions.

Cholesterol in the ciliary membrane as a therapeutic target against cancer.

Primary cilium is a non-motile, antenna-like structure that develops in the quiescent G0 phase-cell surface. It is composed of an array of axonemal microtubules polymerized from the centrosome/basal body. The plasma membrane surrounding the primary cilium, which is called the ciliary membrane, contains a variety of receptors and ion channels, through which the cell receives extracellular chemical and physical stimuli to initiate signal transduction. In general, primary cilia disappear when cells receive the proliferative signals to re-enter the cell cycle. Primary cilia thus cannot be identified in many malignant and proliferative tumors. In contrast, some cancers, including basal cell carcinoma, medulloblastoma, gastrointestinal stromal tumor, and other malignancies, retain their primary cilia. Importantly, it has been reported that the primary cilia-mediated oncogenic signals of Hedgehog, Wnt, and Aurora kinase A are involved in the tumorigenesis and tumor progression of basal cell carcinoma and some types of medulloblastoma. It has also been demonstrated that cholesterol is significantly more enriched in the ciliary membrane than in the rest of the plasma membrane to ensure Sonic hedgehog signaling. A series of epidemiological studies on statin drugs (cholesterol-lowering medication) demonstrated that they prevent recurrence in a wide range of cancers. Taken together, ciliary cholesterol could be a potential therapeutic target in primary cilia-dependent progressive cancers.

High-efficient CRISPR/Cas9-mediated gene targeting to establish cell models of ciliopathies.

Primary cilia are antenna-like structures developed on the cell surface of mammalian cells during the quiescent G0 phase. Primary cilia in mammalian cells receive extracellular signals for early development and cell tissue homeostasis. Ciliopathies characterized with congenital anomalies such as cerebellar hypoplasia, polycystic kidney and polydactyly are caused by germline mutations of ciliary structure- and function-related genes. Gene knock-out techniques in ciliated cultured cells with the uniformed genetic background are useful to evaluate the pathophysiological roles of ciliopathy-related gene products. Genome editing technology has been applied into the gene knock-out in many types of cultured cell lines. However, the frequency of genome editing varies according to cell species and cycle because of dependency on error-free homology-directed repair (HDR) activity. The human telomerase reverse transcriptase-immortalized retinal pigmented epithelial cell line (hTERT-RPE1) is well known for its suitability in cilia research. However, the efficacy of the HDR-mediated knock-out clone isolation was low. Here, we introduce the clustered regularly interspaced short palindromic repeats-obligate ligation-gated recombination (CRISPR-ObLiGaRe) system, which is a nonhomologous end-joining (NHEJ)-mediated gene targeting method, to generate the knock-out clones effectively even in the lower-HDR activity cell lines including hTERT-RPE1 cell. This CRISPR-ObLiGaRe system is a powerful tool for establishing ciliopathy model cell libraries and identifying each gene function in cilia-related phenotypes.

Steroid Rebound Phenomenon as a Cause of Delayed Cerebral Vasospasm in Post-Splenectomy Pneumococcal Meningitis: A Case Report.

Pneumococcal meningitis as an overwhelming post-splenectomy infection (OPSI) has a higher risk of neurological complications and is sometimes life-threatening. In acute pneumococcal meningitis, four days of dexamethasone is widely used for the prevention of neurological complications. Herein, we report a 68-year-old woman with the diagnosis of pneumococcal meningitis as OPSI. With adequate antibiotics and dexamethasone, her symptoms gradually improved. However, after dexamethasone withdrawal, her consciousness got worse and got into a coma. Brain magnetic resonance imaging revealed acute cerebral infarctions in the bilateral middle cerebral artery territory with multiple vascular stenoses and hydrocephalus. Vascular stenoses improved by follow-up, suggesting cerebral vasospasm. There were no suggestive findings of cerebral vasculitis. Follow-up cerebrospinal fluid analysis showed remained pleocytosis with no bacteria, which could not suggest meningitis recurrence. Since steroid therapy was rapidly withdrawn, we diagnosed that the cerebral vasospasm was due to the steroid rebound phenomenon. The steroid rebound phenomenon due to the excessive immune response to bacterial microstructures has been reported in pneumococcal meningitis. Especially, the present case was asplenia and the usual dexamethasone use would not adequately suppress the immune response to bacterial microstructures. Since pneumococcal meningitis as OPSI has a higher risk of neurological complications, clinicians should consider longer and more cautious steroid tapering.

AT-hook DNA-binding motif-containing protein one knockdown downregulates EWS-FLI1 transcriptional activity in Ewing's sarcoma cells.

Ewing's sarcoma is the second most common bone malignancy in children or young adults and is caused by an oncogenic transcription factor by a chromosomal translocation between the EWSR1 gene and the ETS transcription factor family. However, the transcriptional mechanism of EWS-ETS fusion proteins is still unclear. To identify the transcriptional complexes of EWS-ETS fusion transcription factors, we applied a proximal labeling system called BioID in Ewing's sarcoma cells. We identified AHDC1 as a proximal protein of EWS-ETS fusion proteins. AHDC1 knockdown showed a reduced cell growth and transcriptional activity of EWS-FLI1. AHDC1 knockdown also reduced BRD4 and BRG1 protein levels, both known as interacting proteins of EWS-FLI1. Our results suggest that AHDC1 supports cell growth through EWS-FLI1.

Progressive Small Vessel Disease Burden as a Diagnostic of Central Nervous System-Restricted Microscopic Polyangiitis: A Case Report and Review of the Literature.

Microscopic polyangiitis (MPA) is a type of anti-neutrophil cytoplasmic antibody (ANCA)-associated vasculitis linked to myeloperoxidase (MPO), usually accompanied by pulmonary and renal lesions. MPA sometimes causes central nervous system (CNS) involvement such as cerebral infarction. Herein, we report a case of a 72-year-old man with a headache. He had an unknown cause of the elevated inflammatory response. Magnetic resonance imaging (MRI) showed multiple cerebral infarctions in the small vessel region in the right basal ganglia with multiple cerebral microbleeds (cMBs). After admission, his left hemiparesis and consciousness disturbance gradually deteriorated. A follow-up MRI on day 18 showed increased multiple cerebral infarctions in small vessel regions with increased cMBs. Additional blood tests revealed positive MPO-ANCA. Although there were no findings suggestive of active renal or pulmonary involvement or peripheral neuropathy, we diagnosed him as having MPA-associated CNS-restricted vasculitis. CNS involvement of MPA is relatively rare but is associated with a high small vessel disease (SVD) burden. In addition to the unknown cause of inflammatory response, the multiple cMBs increase and a short-term recurrence of cerebral infarctions in the bilateral thalamus and basal ganglia was the clue for the diagnosis of CNS-restricted vasculitis. It is difficult to diagnose MPA vasculitis when lesions are restricted to the CNS. In the absence of lesions other than SVD, MPA-associated CNS vasculitis should be suspected in patients with progressive SVD burden and elevated inflammatory response.

iPSC reprogramming-mediated aneuploidy correction in autosomal trisomy syndromes.

Trisomy 21, 18, and 13 are the major autosomal aneuploidy disorders in humans. They are mostly derived from chromosome non-disjunction in maternal meiosis, and the extra trisomic chromosome can cause several congenital malformations. Various genes on the trisomic chromosomes are intricately involved in the development of disease, and fundamental treatments have not yet been established. However, chromosome therapy has been developed to correct the extra chromosome in cultured patient cells, and it was recently reported that during reprogramming into iPSCs, fibroblasts from a Down syndrome patient lost the extra chromosome 21 due to a phenomenon called trisomy-biased chromosome loss. To gain preliminary insights into the underlying mechanism of trisomy rescue during the early stages of reprogramming, we reprogrammed skin fibroblasts from patients with trisomy syndromes 21, 18, 13, and 9 to iPSC, and evaluated the genomes of the individual iPSC colonies by molecular cytogenetic techniques. We report the spontaneous correction from trisomy to disomy upon cell reprogramming in at least one cell line examined from each of the trisomy syndromes, and three possible combinations of chromosomes were selected in the isogenic trisomy-rescued iPSC clones. Single nucleotide polymorphism analysis showed that the trisomy-rescued clones exhibited either heterodisomy or segmental uniparental isodisomy, ruling out the possibility that two trisomic chromosomes were lost simultaneously and the remaining one was duplicated, suggesting instead that one trisomic chromosome was lost to generate disomic cells. These results demonstrated that trisomy rescue may be a phenomenon with random loss of the extra chromosome and subsequent selection for disomic iPSCs, which is analogous to the karyotype correction in early preimplantation embryos. Our finding is relevant for elucidating the mechanisms of autonomous karyotype correction and future application in basic and clinical research on aneuploidy disorders.

Sulforaphane suppresses metastasis of triple-negative breast cancer cells by targeting the RAF/MEK/ERK pathway.

Breast cancer metastasis is the main cause of cancer death in women, so far, no effective treatment has inhibited breast cancer metastasis. Sulforaphane (SFN), a natural compound derived from broccoli, has shown potential health benefits in many cancers. However, research on breast cancer metastasis is still insufficient. Here, we showed that SFN, including its two isomers of R-SFN and S-SFN, significantly inhibited TGF-ß1-induced migration and invasion in breast cancer cells. Proteomic and phosphoproteomic analysis showed that SFN affected the formation of the cytoskeleton. Subsequent experiments confirmed that SFN significantly inhibited TGF-ß1-induced actin stress fiber formation and the expression of actin stress fiber formation-associated proteins, including paxillin, IQGAP1, FAK, PAK2, and ROCK. Additionally, SFN is directly bound to RAF family proteins (including ARAF, BRAF, and CRAF) and inhibited MEK and ERK phosphorylation. These in vitro results indicate that SFN targets the RAF/MEK/ERK signaling pathway to inhibit the formation of actin stress fibers, thereby inhibiting breast cancer cell metastasis.

Imaging of the Ciliary Cholesterol Underlying the Sonic Hedgehog Signal Transduction.

Primary cilia are antenna-like structures that develop on the surface of quiescent G0-phase cells and receive extracellular signals including sonic hedgehog (Shh) for embryogenesis and adult tissue homeostasis. In mammalian cells, cholesterol activates the seven-transmembrane protein Smoothened to transduce the Shh signal. Germline mutations of the DHCR7 gene encoding the cholesterol biogenesis enzyme 7-dehydrocholesterol reductase cause Smith-Lemli-Opitz syndrome with ciliopathy-related symptoms such as polycystic kidney and polydactyly, implying that cholesterol is indeed involved in ciliary functions. Notably, it has been reported that the cholesterol in ciliary membranes is significantly more abundant than that in the rest of the plasma membrane. However, several studies have failed to image the enriched ciliary cholesterol. Here, we propose a set of protocols for the sensitive imaging of ciliary cholesterol using the fluorescent small compound Filipin III and the green fluorescent protein tagged Domain 4 of the exotoxin Perfringolysin O derived from the anaerobic bacterium Clostridium perfringens. These cholesterol probes should be powerful tools for understanding the physiological and pathological roles of ciliary cholesterol in the context of Shh signaling in mammalian cells.

NBS1 I171V variant underlies individual differences in chromosomal radiosensitivity within human populations.

Genetic information is protected against a variety of genotoxins including ionizing radiation (IR) through the DNA double-strand break (DSB) repair machinery. Genome-wide association studies and clinical sequencing of cancer patients have suggested that a number of variants in the DNA DSB repair genes might underlie individual differences in chromosomal radiosensitivity within human populations. However, the number of established variants that directly affect radiosensitivity is still limited. In this study, we performed whole-exome sequencing of 29 Japanese ovarian cancer patients and detected the NBS1 I171V variant, which is estimated to exist at a rate of approximately 0.15% in healthy human populations, in one patient. To clarify whether this variant indeed contributes to chromosomal radiosensitivity, we generated NBS1 I171V variant homozygous knock-in HCT116 cells and mice using the CRISPR/Cas9 system. Radiation-induced micronucleus formation and chromosomal aberration frequency were significantly increased in both HCT116 cells and mouse embryonic fibroblasts (MEFs) with knock-in of the NBS1 I171V variant compared with the levels in wild-type cells. These results suggested that the NBS1 I171V variant might be a genetic factor underlying individual differences in chromosomal radiosensitivity.

Ciliary GPCR-based transcriptome as a key regulator of cilia length control.

The primary cilium is a plasma membrane-protruding sensory organelle that efficiently conveys signaling cascades in a highly ordered microenvironment. Its signaling is mediated, in part, by a limited set of GPCRs preferentially enriched in the cilium membrane. This includes melanin-concentrating hormone (MCH) receptor 1 (MCHR1), which plays a role in feeding and mood. In addition to its receptor composition, the length of the cilium is a characteristic parameter that is implicated in its function. We previously found that MCH can dynamically shorten cilia length via the Gi/o and Akt pathways in both MCHR1-expressing hTERT-RPE1 cells (hRPE1 cells) and rat hippocampal neurons. However, the detailed mechanisms by which MCH regulates cilia length through ciliary MCHR1 remains unclear. In this study, we aimed to determine the transcriptome changes in MCHR1-expressing hRPE1 cells in response to MCH to identify the target molecules involved in cilia length regulation via MCHR1 activation. RNA sequencing analysis of ciliated cells subjected to MCH treatment showed upregulation of 424 genes and downregulation of 112 genes compared with static control cells. Validation by quantitative real-time PCR, knocking down, and CRISPR/Cas9-mediated knockout technology identified a molecule, PDZ and LIM domain-containing protein 5 (PDLIM5). Thus, it was considered as the most significant key factor for MCHR1-mediated shortening of cilia length. Additional analyses revealed that the actin-binding protein alpha-actinin 1/4 is a crucial downstream target of the PDLIM5 signaling pathway that exerts an effect on MCHR1-induced cilia shortening. In the endogenous MCHR1-expressing hippocampus, transcriptional upregulation of PDLIM5 and actinin 1/4, following the application of MCH, was detected when the MCHR1-positive cilia were shortened. Together, our transcriptome study based on ciliary MCHR1 function uncovered a novel and important regulatory step underlying cilia length control. These results will potentially serve as a basis for understanding the mechanism underlying the development of obesity and mood disorders.