RESUMO
Immune checkpoint inhibitors have significantly transformed the landscape of cancer therapy. Nevertheless, while these inhibitors are highly effective for certain patient groups, many do not benefit due to primary or acquired resistance. Specifically, these treatments often lack sufficient therapeutic efficacy against cancers with low antigenicity. Thus, the development of an effective strategy to overcome cancers with low antigenicity is imperative for advancing next-generation cancer immunotherapy. Here, we show that small molecule inhibitor of hematopoietic progenitor kinase 1 (HPK1) combined with programmed cell death ligand 1 (PD-L1) blockade can enhance T-cell response to tumor with low antigenicity. We found that treatment of OT-1 splenocytes with HPK1 inhibitor enhanced the activation of signaling molecules downstream of T-cell receptor provoked by low-affinity-antigen stimulation. Using an in vivo OT-1 T-cell transfer model, we demonstrated that combining the HPK1 inhibitor with the anti-PD-L1 antibody significantly suppressed the growth of tumors expressing low-affinity altered peptide ligand of chicken ovalbumin, while anti-PD-L1 antibody monotherapy was ineffective. Our findings offer crucial insights into the potential for overcoming tumors with low antigenicity by combining conventional immune checkpoint inhibitors with HPK1 inhibitor.
Assuntos
Antígeno B7-H1 , Camundongos Endogâmicos C57BL , Proteínas Serina-Treonina Quinases , Animais , Antígeno B7-H1/antagonistas & inibidores , Antígeno B7-H1/imunologia , Camundongos , Proteínas Serina-Treonina Quinases/antagonistas & inibidores , Proteínas Serina-Treonina Quinases/imunologia , Proteínas Serina-Treonina Quinases/metabolismo , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Linhagem Celular Tumoral , Linfócitos T/imunologia , Linfócitos T/efeitos dos fármacos , Imunoterapia/métodos , Inibidores de Proteínas Quinases/farmacologia , Inibidores de Proteínas Quinases/uso terapêutico , Humanos , Neoplasias/imunologia , Neoplasias/tratamento farmacológico , Neoplasias/patologia , Proteínas Quinases Dependentes de 3-FosfoinositídeoRESUMO
Fibrolamellar hepatocellular carcinoma (FL-HCC) is known as a highly aggressive liver cancer that typically affects young adults without virus infection. Since this type of cancer does not respond to chemotherapy, surgery is the only known effective therapeutic option. Most FL-HCC patients express the fusion gene DNAJB1-PRKACA, which has been recognized as the signature of FL-HCC. It has also been reported that PRKACA kinase activity is essential for its oncogenic activity, suggesting that PRKACA kinase inhibition could be considered as an useful therapeutic target. In this study, we established an evaluation system for PRKACA kinase inhibitors and synthesized DS89002333, a novel PRKACA inhibitor. DS89002333 showed potent PRKACA inhibitory activity and inhibited fusion protein-dependent cell growth both in vitro and in vivo. Furthermore, this compound showed anti-tumor activity in an FL-HCC patient-derived xenograft model expressing the DNAJB1-PRKACA fusion gene. Our data suggest that DS89002333 could be considered as a potential therapeutic agent for FL-HCC.
Assuntos
Carcinoma Hepatocelular , Neoplasias Hepáticas , Carcinoma Hepatocelular/tratamento farmacológico , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/metabolismo , Domínio Catalítico , Subunidades Catalíticas da Proteína Quinase Dependente de AMP Cíclico/genética , Subunidades Catalíticas da Proteína Quinase Dependente de AMP Cíclico/metabolismo , Regulação Neoplásica da Expressão Gênica , Proteínas de Choque Térmico HSP40/genética , Proteínas de Choque Térmico HSP40/metabolismo , Humanos , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/metabolismo , Inibidores de Proteínas Quinases , Adulto JovemRESUMO
BACKGROUND: Although increasing evidence indicates that even variations in normal range thyroid function are associated with Alzheimer's disease (AD), the association between serum thyroid hormone levels within the reference range and AD biomarkers remains unclear. This study examined whether variations in thyroid hormones within the reference range are associated with brain amyloid burden and cortical glucose metabolism in older adults without dementia. METHODS: One hundred and two non-demented older adults underwent 11 C-Pittsburgh Compound B positron emission tomography (PiB-PET), 18 F-fluorodeoxyglucose (FDG)-PET, and measurement of serum thyroid-stimulating hormone (TSH), free triiodothyronine (T3), and free thyroxine (T4) levels. The discrimination between PiB-negative and PiB-positive subgroup was made on the basis of a subject's cortical uptake value ratio greater than 1.4. The association of serum thyroid hormone levels with global PiB or FDG uptake, and PiB or FDG uptake in each region of interest, including frontal and temporoparietal lobes and posterior cingulate gyrus, was analysed using a multiple regression model with adjustment for covariates, including age, gender, years of education, apolipoprotein E4 status or PiB uptake value. RESULTS: In the PiB-positive subgroup, the serum TSH levels positively associated with the global FDG uptake (ß = 0.471, P = 0.003) and FDG uptake in the frontal and temporoparietal lobes (ß = 0.466, P = 0.003, ß = 0.394, P = 0.012, respectively); the serum-free T3 levels negatively associated with the FDG uptake in the temporoparietal lobe and posterior cingulate region (ß = -0.351, P = 0.033, ß = -0.544, P = 0.002, respectively). The PiB-negative subgroup showed no significant associations. The serum thyroid hormone levels did not correlate with the global PiB uptake and PiB uptake in each region. CONCLUSIONS: The variations in the thyroid hormones within the reference ranges are associated with glucose metabolism, particularly in the specific regions affected by the neuropathologic changes of AD, in non-demented older adults with brain amyloid burden.
Assuntos
Doença de Alzheimer , Fluordesoxiglucose F18 , Idoso , Doença de Alzheimer/metabolismo , Amiloide/metabolismo , Encéfalo/diagnóstico por imagem , Encéfalo/metabolismo , Fluordesoxiglucose F18/metabolismo , Glucose/metabolismo , Humanos , Tomografia por Emissão de Pósitrons/métodos , Glândula Tireoide/diagnóstico por imagem , Glândula Tireoide/metabolismo , Hormônios Tireóideos/metabolismo , Tireotropina/metabolismo , Tomografia Computadorizada por Raios XRESUMO
A series of imidazolinylindole derivatives were discovered as novel kallikrein 7 (KLK7, stratum corneum chymotryptic enzyme) inhibitors. Structure-activity relationship (SAR) studies led to the identification of potent human KLK7 inhibitors. By further modification of the benzenesulfonyl moiety to overcome species differences in inhibitory activity, potent inhibitors against both human and mouse KLK7 were identified. Furthermore, the complex structure of 25 with mouse KLK7 could explain the SAR and the cause of the species differences in inhibitory activity.
Assuntos
Descoberta de Drogas , Inibidores Enzimáticos/farmacologia , Imidazolinas/farmacologia , Indóis/farmacologia , Calicreínas/antagonistas & inibidores , Animais , Relação Dose-Resposta a Droga , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Humanos , Imidazolinas/síntese química , Imidazolinas/química , Indóis/síntese química , Indóis/química , Calicreínas/metabolismo , Camundongos , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
The protective effects of G protein-coupled receptor 39 (GPR39) on concanavalin A (Con A)-induced hepatitis in mice was examined. In a dose dependent manner and at 24 h after the elicitation by Con A, oral administration of TC-G 1008, a GPR39 agonist, reduced both, the glutamic-pyruvic transaminase levels (a marker for liver injury) and the necrosis area, as revealed by the histological analysis of tissues from mice with Con A-induced hepatitis. TC-G 1008 also suppressed serum interleukin (IL)-6 and tumor necrosis factor (TNF)-α significantly at 6 h after the elicitation, suggesting that the cells producing IL-6 and/or TNF-α are the targets of TC-G 1008. One potential target cell appears to be a monocyte-derived macrophages because TC-G 1008 treatment suppressed lipopolysaccharide-induced IL-6 production from U937 macrophages in vitro. Taken together, GPR39 agonist TC-G 1008 ameliorates liver injury in the Con A model by blocking pro-inflammatory cytokine production. Use of GPR39 agonists for monotherapy or in combination with immunosuppressants might prove to be beneficial in the treatment of autoimmune hepatitis.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas/tratamento farmacológico , Hepatite/tratamento farmacológico , Pirimidinas/farmacologia , Receptores Acoplados a Proteínas G/agonistas , Receptores Acoplados a Proteínas G/metabolismo , Sulfonamidas/farmacologia , Animais , Técnicas de Cultura de Células , Doença Hepática Induzida por Substâncias e Drogas/patologia , Concanavalina A/farmacologia , Humanos , Imidazóis/farmacologia , Interleucina-10/sangue , Interleucina-10/metabolismo , Interleucina-6/sangue , Interleucina-6/metabolismo , Fígado/efeitos dos fármacos , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Modelos Animais , Pirazóis/farmacologia , Piridazinas/farmacologia , Fator de Necrose Tumoral alfa/sangue , Fator de Necrose Tumoral alfa/metabolismo , Células U937RESUMO
A novel series of 1,3,6-trisubstituted 1,4-diazepan-7-ones were investigated as human kallikrein 7 (KLK7, stratum corneum chymotryptic enzyme) inhibitors. Based on the X-ray co-crystal structure of compound 1 bound to human KLK7, the derivatives of this scaffold were designed, synthesized, and evaluated. Through structure-activity relationship studies focused on the side chain located in the prime site region of the enzyme, representative compounds 15, 33a, and 35a were identified as highly potent and selective inhibitors of human KLK7.
Assuntos
Azepinas/farmacologia , Calicreínas/antagonistas & inibidores , Azepinas/síntese química , Azepinas/química , Sítios de Ligação , Cristalografia por Raios X , Desenho de Fármacos , Humanos , Calicreínas/química , Simulação de Acoplamento Molecular , Estrutura Molecular , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
A series of 1,3,6-trisubstituted 1,4-diazepan-7-ones were prepared as kallikrein 7 (KLK7, stratum corneum chymotryptic enzyme) inhibitors. Previously reported compounds 1-3 were potent human KLK7 inhibitors; however, they did not exhibit inhibitory activity against mouse KLK7. Comparison of the human and mouse KLK7 structures reveals the cause of this species differences; therefore, compounds that could inhibit both KLK7s were designed, synthesized, and evaluated. Through this structure-based drug design, compound 22g was identified as an inhibitor against human and mouse KLK7, and only one of the enantiomers, (-)-22g, exhibited potent inhibitory activity. Furthermore, the crystal structure of mouse KLK7 complexed with 22g enabled the elucidation of structure-activity relationships and justified 22g as a valuable compound to overcome the species differences.
Assuntos
Azepinas/química , Calicreínas/metabolismo , Inibidores de Proteases/síntese química , Sequência de Aminoácidos , Animais , Azepinas/metabolismo , Sítios de Ligação , Cristalografia por Raios X , Desenho de Fármacos , Humanos , Calicreínas/antagonistas & inibidores , Camundongos , Inibidores de Proteases/metabolismo , Estrutura Terciária de Proteína , Alinhamento de Sequência , Especificidade da Espécie , Estereoisomerismo , Relação Estrutura-AtividadeRESUMO
Compound 1, composed of a 1,3,6-trisubstituted 1,4-diazepane-7-one, was discovered as a novel human kallikrein 7 (KLK7, stratum corneum chymotryptic enzyme, SCCE) inhibitor, and its derivatives were synthesized and evaluated. Structure-activity relationship studies of the amidoxime unit and benzoic acid part of this new scaffold led to the identification of 25 and 34, which were more potent than the hit compound, 1. The X-ray co-crystal structure of compound 25 and human KLK7 revealed the characteristic interactions and enabled explanations of the structure-activity relationship.
Assuntos
Azepinas/farmacologia , Descoberta de Drogas , Calicreínas/antagonistas & inibidores , Inibidores de Serina Proteinase/farmacologia , Azepinas/síntese química , Azepinas/química , Relação Dose-Resposta a Droga , Humanos , Calicreínas/metabolismo , Estrutura Molecular , Inibidores de Serina Proteinase/síntese química , Inibidores de Serina Proteinase/química , Relação Estrutura-AtividadeRESUMO
BACKGROUNDPrecise stratification of patients with non-small cell lung cancer (NSCLC) is needed for appropriate application of PD-1/PD-L1 blockade therapy.METHODSWe measured soluble forms of the immune-checkpoint molecules PD-L1, PD-1, and CTLA-4 in plasma of patients with advanced NSCLC before PD-1/PD-L1 blockade. A prospective biomarker-finding trial (cohort A) included 50 previously treated patients who received nivolumab. A retrospective observational study was performed for patients treated with any PD-1/PD-L1 blockade therapy (cohorts B and C), cytotoxic chemotherapy (cohort D), or targeted therapy (cohort E). Plasma samples from all patients were assayed for soluble immune-checkpoint molecules with a highly sensitive chemiluminescence-based assay.RESULTSNonresponsiveness to PD-1/PD-L1 blockade therapy was associated with higher concentrations of these soluble immune factors among patients with immune-reactive (hot) tumors. Such an association was not apparent for patients treated with cytotoxic chemotherapy or targeted therapy. Integrative analysis of tumor size, PD-L1 expression in tumor tissue (tPD-L1), and gene expression in tumor tissue and peripheral CD8+ T cells revealed that high concentrations of the 3 soluble immune factors were associated with hyper or terminal exhaustion of antitumor immunity. The combination of soluble PD-L1 (sPD-L1) and sCTLA-4 efficiently discriminated responsiveness to PD-1/PD-L1 blockade among patients with immune-reactive tumors.CONCLUSIONCombinations of soluble immune factors might be able to identify patients unlikely to respond to PD-1/PD-L1 blockade as a result of terminal exhaustion of antitumor immunity. Our data suggest that such a combination better predicts, along with tPD-L1, for the response of patients with NSCLC.TRIAL REGISTRATIONUMIN000019674.FUNDINGThis study was funded by Ono Pharmaceutical Co. Ltd. and Sysmex Corporation.
Assuntos
Carcinoma Pulmonar de Células não Pequenas , Inibidores de Checkpoint Imunológico , Neoplasias Pulmonares , Humanos , Antígeno B7-H1 , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Carcinoma Pulmonar de Células não Pequenas/genética , Carcinoma Pulmonar de Células não Pequenas/patologia , Fatores Imunológicos/sangue , Fatores Imunológicos/química , Neoplasias Pulmonares/tratamento farmacológico , Neoplasias Pulmonares/genética , Neoplasias Pulmonares/patologia , Receptor de Morte Celular Programada 1 , Inibidores de Checkpoint Imunológico/farmacologia , Inibidores de Checkpoint Imunológico/uso terapêuticoRESUMO
Duchenne muscular dystrophy (DMD) is an X-linked fatal muscular disease, affecting one in 3,500 live male births worldwide. Currently, there is no cure for this disease, except for steroid-based treatment to attenuate disease progression. Cell transplantation therapy is a promising therapeutic approach, however, there is a lack of appropriate animal models to conduct large-scale preclinical studies using human cells, including biochemical and functional tests. Here, we established an immunodeficient DMD rat model and performed exhaustive pathological analysis and transplantation efficiency evaluation to assess its suitability to study DMD. Our DMD rat model exhibited histopathological characteristics similar to those observed in human patients with DMD. Human myoblasts demonstrated successful engraftment following transplantation into these rats. Therefore, this immunodeficient DMD rat model would be useful in preclinical studies to develop cellular transplantation therapies for DMD.
RESUMO
Augmenting T-cell activity is a promising approach to enhance the efficacy of cancer immunotherapy treatment. Hematopoietic progenitor kinase 1 (HPK1) is predominantly expressed in immune cells and negatively regulates T-cell receptor signaling. It is reported that inhibition of the kinase function of HPK1 results in tumor growth suppression by enhancing cancer immunity. Thus, developing HPK1 inhibitors has attracted considerable attention as a future cancer immunotherapy approach. However, despite recent progress in HPK1 biology and pharmacology, various challenges still remain, such as developing HPK1 inhibitors with favorable pharmacological profiles and identifying tumor characteristics that can be applied to define susceptibility to HPK1 inhibition. Here, we present the identification and pharmacological evaluation of DS21150768, a potent small-molecule HPK1 inhibitor with a novel chemical scaffold. DS21150768 shows remarkable inhibition of HPK1 kinase activity, and in vitro studies demonstrated its potent activity to enhance T-cell function. DS21150768 is orally bioavailable and shows sustained plasma exposure, which leads to enhanced cytokine responses in vivo. We conducted a comparison of the anti-tumor efficacy of DS21150768 alone or in combination with anti-PD-1 antibody in 12 different mouse cancer cell models, and observed that the treatments suppressed tumor growth in multiple models. Furthermore, Gene Set Enrichment Analysis demonstrated significant enrichment of immune-related gene signatures in the tumor models responsive to DS21150768 treatment. Our results provide a path forward for the future development of HPK1 inhibitors and fundamental insights into biomarkers of HPK1-targeted therapy.
Assuntos
Neoplasias , Camundongos , Animais , Neoplasias/tratamento farmacológico , Linfócitos T , Transdução de Sinais , CitocinasRESUMO
Histone acetylation is a post-translational modification of histones that is catalyzed by histone acetyltransferases (HATs) and plays an essential role in cellular processes. The HAT domain of EP300/CBP has recently emerged as a potential drug target for cancer therapy. Here, we describe the identification of the novel, highly potent, and selective EP300/CBP HAT inhibitor DS-9300. Our optimization efforts using a structure-based drug design approach based on the cocrystal structures of the EP300 HAT domain in complex with compounds 2 and 3 led to the identification of compounds possessing low-nanomolar EP300 HAT inhibitory potency and the ability to inhibit cellular acetylation of histone H3K27. Optimization of the pharmacokinetic properties in this series resulted in compounds with excellent oral systemic exposure, and once-daily oral administration of 16 (DS-9300) demonstrated potent antitumor effects in a castrated VCaP xenograft mouse model without significant body weight loss.
Assuntos
Histona Acetiltransferases , Histonas , Humanos , Camundongos , Animais , Histonas/metabolismo , Histona Acetiltransferases/metabolismo , Acetilação , Fatores de Transcrição de p300-CBP , Proteína p300 Associada a E1ARESUMO
A series of truncated analogs of α-galactosylceramide with altered ceramide moiety was prepared, and evaluated for Th2-biased response in the context of IL-4/IFN-γ ratio. Phytosphingosine-modified analogs including cyclic, aromatic and ethereal compounds as well as the C-glycoside analog of OCH (2) with their cytokine inducing profile are disclosed.
Assuntos
Galactosilceramidas/química , Galactosilceramidas/farmacologia , Regulação da Expressão Gênica/efeitos dos fármacos , Interferon gama/metabolismo , Interleucina-4/metabolismo , Animais , Antígenos CD1d/química , Antígenos CD1d/metabolismo , Sítios de Ligação , Simulação por Computador , Galactosilceramidas/síntese química , Camundongos , Camundongos Endogâmicos C57BL , Células Th2/efeitos dos fármacosRESUMO
The 2009 pandemic H1N1 influenza A virus spread quickly worldwide in 2009. Since most of the fatal cases were reported in developing countries, rapid and accurate diagnosis methods that are usable in poorly equipped laboratories are necessary. In this study, a mobile detection system for the 2009 H1N1 influenza A virus was developed using a reverse-transcriptase loop-mediated isothermal amplification (RT-LAMP) kit with a disposable pocket-warmer as a heating device (designated as pwRT-LAMP). The pwRT-LAMP can detect as few as 100 copies of the virus--which is nearly as sensitive as real-time reverse-transcription polymerase chain reaction (RT-PCR)--and does not cross-react with RNA of seasonal influenza viruses. To evaluate the usefulness of the pwRT-LAMP system, nasal swab samples were collected from 56 patients with flu-like symptoms and were tested. Real-time RT-PCR confirmed that the 2009 H1N1 influenza A virus was present in 27 of the 56 samples. Of these 27 positive samples, QuickVue Influenza A+B immunochromatography detected the virus in only 11 samples (11/27; 40.7%), whereas the pwRT-LAMP system detected the virus in 26 of the 56 samples (26/27 of the positive samples; 96.3%). These findings indicate that the mobile pwRT-LAMP system is an accurate diagnostic system for the 2009 H1N1 influenza A virus, and has great potential utility in diagnosing future influenza pandemics.
Assuntos
Vírus da Influenza A Subtipo H1N1/isolamento & purificação , Influenza Humana/diagnóstico , Influenza Humana/virologia , Técnicas de Amplificação de Ácido Nucleico/métodos , Sistemas Automatizados de Assistência Junto ao Leito , Virologia/métodos , Adulto , Feminino , Humanos , Vírus da Influenza A Subtipo H1N1/genética , Masculino , Pessoa de Meia-Idade , Sensibilidade e EspecificidadeRESUMO
Collagen VI is distributed in the interstitium and is secreted mainly by mesenchymal stromal cells (MSCs) in skeletal muscle. Mutations in COL6A1-3 genes cause a spectrum of COL6-related myopathies. In this study, we performed a systemic transplantation study of human-induced pluripotent stem cell (iPSC)-derived MSCs (iMSCs) into neonatal immunodeficient COL6-related myopathy model (Col6a1 KO /NSG) mice to validate the therapeutic potential. Engraftment of the donor cells and the resulting rescued collagen VI were observed at the quadriceps and diaphragm after intraperitoneal iMSC transplantation. Transplanted mice showed improvement in pathophysiological characteristics compared with untreated Col6a1 KO /NSG mice. In detail, higher muscle regeneration in the transplanted mice resulted in increased muscle weight and enlarged myofibers. Eight-week-old mice showed increased muscle force and performed better in the grip and rotarod tests. Overall, these findings support the concept that systemic iMSC transplantation can be a therapeutic option for COL6-related myopathies.
RESUMO
The effects of amino acid variants encoded by human leukocyte antigen (HLA) class II on the development of Graves' disease (GD) and Hashimoto's thyroiditis (HT) have not been fully elucidated. We investigated the HLA-DRB1 genes of 243 GD patients and 82 HT patients in the Japanese population and compared the frequencies of HLA-DRB1 alleles and HLA-DRB1 amino acid variants between these patients and the Japanese populations previously reported by another institution. The frequencies of HLA-DRB1*04:05 and -DRB1*14:03 alleles were significantly higher and those of HLA-DRB1*01:01 and -DRB1*15:02 alleles were lower in GD patients than in controls. The frequencies of HLA-DRB1*08:03 and -DRB1*09:01 alleles were significantly higher and that of the HLA-DRB1*13:02 allele was lower in HT patients than in controls. A blind association analysis with all amino acid positions identified DRß9 and DRß31 for GD and DRß9, DRß13, and DRß21 for HT. The frequency of Glu-9 was significantly higher and that of Cys-9 was lower in GD patients than in controls. The frequencies of Lys-9 and Phe-13 were significantly higher in HT patients than in controls. DRß9 and DRß13 could be critical amino acid positions in the development of GD and HT.
Assuntos
Aminoácidos/genética , Genótipo , Doença de Graves/imunologia , Cadeias HLA-DRB1/genética , Doença de Hashimoto/imunologia , Adulto , Idoso , Alelos , Feminino , Frequência do Gene , Estudos de Associação Genética , Predisposição Genética para Doença , Doença de Graves/genética , Doença de Hashimoto/genética , Humanos , Japão , Masculino , Pessoa de Meia-Idade , Polimorfismo GenéticoRESUMO
BACKGROUND: Mesenchymal stromal cells (MSCs) function as supportive cells on skeletal muscle homeostasis through several secretory factors including type 6 collagen (COL6). Several mutations of COL6A1, 2, and 3 genes cause Ullrich congenital muscular dystrophy (UCMD). Skeletal muscle regeneration deficiency has been reported as a characteristic phenotype in muscle biopsy samples of human UCMD patients and UCMD model mice. However, little is known about the COL6-dependent mechanism for the occurrence and progression of the deficiency. The purpose of this study was to clarify the pathological mechanism of UCMD by supplementing COL6 through cell transplantation. METHODS: To test whether COL6 supplementation has a therapeutic effect for UCMD, in vivo and in vitro experiments were conducted using four types of MSCs: (1) healthy donors derived-primary MSCs (pMSCs), (2) MSCs derived from healthy donor induced pluripotent stem cell (iMSCs), (3) COL6-knockout iMSCs (COL6KO-iMSCs), and (4) UCMD patient-derived iMSCs (UCMD-iMSCs). RESULTS: All four MSC types could engraft for at least 12 weeks when transplanted into the tibialis anterior muscles of immunodeficient UCMD model (Col6a1KO) mice. COL6 protein was restored by the MSC transplantation if the MSCs were not COL6-deficient (types 1 and 2). Moreover, muscle regeneration and maturation in Col6a1KO mice were promoted with the transplantation of the COL6-producing MSCs only in the region supplemented with COL6. Skeletal muscle satellite cells derived from UCMD model mice (Col6a1KO-MuSCs) co-cultured with type 1 or 2 MSCs showed improved proliferation, differentiation, and maturation, whereas those co-cultured with type 3 or 4 MSCs did not. CONCLUSIONS: These findings indicate that COL6 supplementation improves muscle regeneration and maturation in UCMD model mice.
Assuntos
Colágeno Tipo VI , Músculo Esquelético , Animais , Transplante de Células , Colágeno Tipo VI/genética , Suplementos Nutricionais , Humanos , Camundongos , Distrofias Musculares , EscleroseRESUMO
We studied the reflex actions of the cutaneous afferents innervating the trunk to hindlimb motoneurons in the spinal cat using an intracellular recording technique. Stimulation of the trunk cutaneous afferents entering into the L2-L5 spinal segment produced different types of polysynaptic potentials in hindlimb motoneurons via polysynaptic neuronal pathways. The trunk cutaneous afferents predominantly caused excitatory PSPs in the flexor motoneurons and inhibitory PSPs in the extensor motoneurons. The size and latency of polysynaptic potentials were related to the proximity of the spinal segments of the nerves stimulated to the spinal segments of motoneurons. These findings suggest that the neuronal pathways from trunk cutaneous afferents to hindlimb motoneurons play an important role in coordinating between the trunk and hindlimbs.
Assuntos
Gatos/fisiologia , Neurônios Motores/fisiologia , Músculo Esquelético/inervação , Vias Neurais/fisiologia , Medula Espinal/fisiologia , Vias Aferentes , Animais , Eletrofisiologia , Potenciais Pós-Sinápticos Excitadores , Feminino , Membro Posterior/fisiologia , Masculino , Inibição NeuralRESUMO
The three-dimensional distribution of dendrites from motoneurons innervating longissimus lumborum (Long Motoneurons) in the L4 spinal segment was examined in the adult cat using intracellular staining techniques. Long Motoneurons were electrophysiologically identified, stained with injection of biocytin and reconstructed from serial histological sections. Somas of Long Motoneurons were mainly located in the lateral-ventral area of the ventral horn. The dendritic distribution followed an orderly pattern in all motoneurons examined. Long Motoneurons showed a multi-directional distribution of dendrites, and the dendritic distribution pattern varied depending on the motoneuron. All studied motoneurons distributed dendrites from the spine into the white matter. The most significant morphological characteristic of the Long Motoneurons was the variation in their dendritic distribution. No relationship was observed between the effects of peripheral afferent inputs from the hindlimb and morphological characteristics of motoneurons.
Assuntos
Dendritos/ultraestrutura , Neurônios Motores/citologia , Animais , Gatos , Corantes , Estado de Descerebração/patologia , Estado de Descerebração/veterinária , Feminino , Membro Posterior/inervação , Lisina/análogos & derivados , Masculino , Músculo Esquelético/inervaçãoRESUMO
Natural killer T (NKT) cells are known to produce Th17 cytokine IL-17 in addition to Th1/2 cytokines. In this study, the ability of NKT cells to produce IL-22, another Th17 cytokine, was examined in mice. When murine spleen cells were stimulated with alpha-galactosyl ceramide, a ligand for NKT cells, not only Th1/2 cytokines (IFN-gamma, IL-4) but Th17 cytokines (IL-17, IL-22) were produced. NKT cells isolated from splenocytes released IL-17 and IL-22 following CD3, CD3/IL-2 or CD3/CD28 stimulation, in which CD3/CD28 costimulation was most effective. Production of IL-17 and IL-22 in CD4+ and CD8+ T cells from splenocytes was little, if any, even after CD3/CD28 costimulation. Treatment with IL-6/TGF-beta decreased CD3/CD28-stimulated production of IL-22, but not that of IL-17, in NKT cells. These findings show for the first time that NKT cells are a cell source of IL-22, and that expression of two Th17 cytokines might be regulated in NKT cells by different mechanisms.