RESUMEN
Dengue virus (DENV) infection is a public health concern in several countries and is associated with severe diseases, such as dengue hemorrhagic fever and dengue shock syndrome. DENVs are transmitted to humans via the bites of infected Aedes mosquitoes, and no antiviral therapeutics are currently available. In this work, we aimed to identify antiviral drugs against DENV type 2 (DENV2) infections and selected pimecrolimus as a potential antiviral drug candidate. Pimecrolimus significantly inhibited DENV2-mediated cell death and replication in vitro. We also confirmed a decrease in the number of plaques formed as well as in the envelope protein levels of DENV2. The time-of-addition and course experiments revealed that pimecrolimus inhibited DENV2 infection during the early stages of the virus replication cycle. In an experimental mouse model, orally administered pimecrolimus alleviated body weight loss and lethality caused by DENV2 infection, which we used as readouts of the drug's antiviral potency. Furthermore, pimecrolimus significantly inhibited the DENV2 load and ameliorated focal necrosis in the liver and spleen. Taken together, our in vitro and in vivo findings suggest that pimecrolimus is a promising antiviral drug candidate for the treatment of DENV2 infection.
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Antivirales , Virus del Dengue , Dengue , Tacrolimus , Replicación Viral , Animales , Virus del Dengue/efectos de los fármacos , Antivirales/farmacología , Antivirales/uso terapéutico , Tacrolimus/análogos & derivados , Tacrolimus/farmacología , Tacrolimus/uso terapéutico , Replicación Viral/efectos de los fármacos , Ratones , Dengue/tratamiento farmacológico , Dengue/virología , Humanos , Modelos Animales de Enfermedad , Chlorocebus aethiops , Línea Celular , Células VeroRESUMEN
Antascomicin B is a natural product that similarly to the macrolides FK506 and Rapamycin binds to the FK506-binding protein 12 (FKBP12). FK506 and Rapamycin act as molecular glues by inducing ternary complexes between FKBPs and additional target proteins. Whether Antascomicin B can induce ternary complexes is unknown. Here we show that Antascomicin B binds tightly to larger human FKBP homologs. The cocrystal structure of FKBP51 in complex with Antascomicin B revealed that large parts of Antascomicin B are solvent-exposed and available to engage additional proteins. Cellular studies demonstrated that Antascomicin B enhances the interaction between human FKBP51 and human Akt. Our studies show that molecules with molecular glue-like properties are more prominent in nature than previously thought. We predict the existence of additional 'orphan' molecular glues that evolved to induce ternary protein complexes but where the relevant ternary complex partners are unknown.
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Proteínas Proto-Oncogénicas c-akt , Proteínas de Unión a Tacrolimus , Tacrolimus , Humanos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Sirolimus/farmacología , Tacrolimus/farmacología , Tacrolimus/análogos & derivados , Proteínas de Unión a Tacrolimus/química , Proteínas de Unión a Tacrolimus/metabolismoRESUMEN
Calcineurin inhibitors have been found to exhibit a preventive role against neuroinflammation, which represents a crucial underlying mechanism in neurodegenerative diseases (ND). Additionally, they possess suppressive effects on the activation of apoptotic pathways, which constitute another mechanism underlying such diseases. Given that pimecrolimus, a calcineurin inhibitor, impedes the synthesis of pro-inflammatory cytokines, such as interleukin (IL)-2, IL-4, and IL-10, and influences apoptotic processes, it is noteworthy to test its potential neuroprotective properties. Thus, the objective of this investigation was to assess the potential protective effects of pimecrolimus against the degenerative consequences of both microglial secretomes and hydrogen peroxide (H2O2), an oxidant agent. The survival rates of HMC3 microglia cells, neuron-like differentiated SH-SY5Y (d-SH-SY5Y) cells, and their co-culture were determined using the 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyltetrazolium Bromide (MTT) method. Furthermore, the levels of pro-inflammatory cytokines IL-1ß and IL-6, and anti-inflammatory cytokine IL-10 were measured using ELISA kits, besides total antioxidant and oxidant capacities in conditioned media of cells. Additionally, the effect of pimecrolimus on neurite length in these cell groups was evaluated through morphological observations. This study revealed, for the first time, that pimecrolimus exerts preventive effects on neurodegenerative processes by virtue of its anti-inflammatory and -antioxidant activities. It holds promise as a potential treatment option for ND.
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Antioxidantes , Neuroblastoma , Tacrolimus/análogos & derivados , Humanos , Antioxidantes/farmacología , Peróxido de Hidrógeno , Interleucina-10 , Microglía , Secretoma , Neuronas , Oxidantes , Citocinas , Antiinflamatorios/farmacologíaRESUMEN
INTRODUCTION: Atopic dermatitis (AD) exhibits difference in immune polarization between Caucasians and Asian races due to which an evaluation of the efficacy and safety of Pimecrolimus (PIM) in Asian population is called for. The current study addresses the need via a sub-group analysis of the PETITE study (NCT00120523) to evaluate the safety and efficacy of PIM in Chinese infants. MATERIALS AND METHODS: Patients with AD (≥3 months-<12 months of age) were randomized in a 1:1 ratio to either PIM 1% cream or topical corticosteroids (TCS). The primary endpoint was safety. The secondary endpoint was efficacy. RESULTS: 120 patients were randomized to either PIM 1% or TCS (n = 61 for PIM, n = 59 for TCS). The most often reported adverse events were reported by similar proportions of patients treated with PIM or TCS. There was a progressive increase in overall IGA treatment success in infants treated with PIM (82.9%, p < .05, 95% CI: 70.4, 95.3) after 26 weeks which was comparable to the TCS group (88.5%, p < .05, 95% CI: 79.8, 97.1). CONCLUSION: PIM showed an early and sustained efficacy in the Chinese sub-population with a substantial corticosteroid-sparing effect in patients with AD.
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Dermatitis Atópica , Fármacos Dermatológicos , Tacrolimus , Humanos , Lactante , Dermatitis Atópica/tratamiento farmacológico , Fármacos Dermatológicos/administración & dosificación , Fármacos Dermatológicos/efectos adversos , Fármacos Dermatológicos/uso terapéutico , Pueblos del Este de Asia , Tacrolimus/administración & dosificación , Tacrolimus/efectos adversos , Tacrolimus/análogos & derivados , Tacrolimus/uso terapéutico , Resultado del Tratamiento , Administración Tópica , Glucocorticoides/administración & dosificación , Glucocorticoides/efectos adversos , Glucocorticoides/uso terapéutico , Crema para la PielRESUMEN
BACKGROUND/AIM: Co-treatment with calcineurin inhibitors, such as tacrolimus and cyclosporin A, can sensitize chemotherapy-resistant cancer cells with P-glycoprotein (P-gp)-over-expression. Pimecrolimus (PIME) is a clinically available calcineurin inhibitor with a structure similar to that of tacrolimus. Whether PIME can sensitize P-gp-over-expressing resistant cancer cells remains unclear. MATERIALS AND METHODS: Cell viability assay, annexin V analyses, cellular morphology and density observation with a microscope, western-blotting, fluorescence-activated cell sorting (FACS), and analysis for P-gp inhibitory activity were performed to investigate the mechanism of action. RESULTS: PIME exhibited strong cytotoxicity to vincristine (VIC)-treated drug-resistant cell lines (KBV20C and MCF-7/ADR) over-expressing P-gp. Co-treatment with VIC and PIME increased apoptosis and down-regulated the ERK signaling pathway, resulting in G2 arrest. PIME could be co-administered with vinorelbine or eribulin to sensitize resistant KBV20C or MCF-7/ADR cancer cells. Moreover, PIME strongly inhibited the efflux of both rhodamine 123 and calcein-AM substrates through P-gp after 4 h of treatment, indicating that VIC+PIME sensitized cancer cells by inhibiting VIC efflux via direct PIME binding to P-gp. Low doses of PIME, tacrolimus, and cyclosporin A showed similar sensitizing efficiencies in resistant KBV20C cells. These drugs showed similar P-gp inhibitory activities using both rhodamine 123 and calcein-AM substrates, suggesting that calcineurin inhibitors generally have strong P-gp inhibitory activities that sensitize drug-resistant cancer cells with P-gp over-expression. CONCLUSION: PIME, currently used in clinics, can be repositioned for treating patients with P-gp-over-expressing resistant cancer (stem) cells.
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Miembro 1 de la Subfamilia B de Casetes de Unión a ATP , Inhibidores de la Calcineurina , Neoplasias , Tacrolimus , Humanos , Subfamilia B de Transportador de Casetes de Unión a ATP/antagonistas & inhibidores , Subfamilia B de Transportador de Casetes de Unión a ATP/efectos de los fármacos , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/antagonistas & inhibidores , Miembro 1 de la Subfamilia B de Casetes de Unión a ATP/efectos de los fármacos , Inhibidores de la Calcineurina/farmacología , Ciclosporina/farmacología , Rodamina 123 , Tacrolimus/análogos & derivados , Tacrolimus/farmacologíaRESUMEN
BACKGROUND: The calcineurin pathway is often activated in mycosis fungoides. We aimed to assess the activity and safety of topical pimecrolimus, a calcineurin inhibitor, in patients with early mycosis fungoides. METHODS: PimTo-MF was a single-arm, multicentre, phase 2 trial done at six medical centres in Spain. Patients (aged ≥18 years) had histologically confirmed early mycosis fungoides (stages IA-IIA) and an Eastern Cooperative Oncology Group performance status of 0-1. Key exclusion criteria included the use of concurrent treatments for mycosis fungoides, including sunbathing, topical or systemic corticosteroids, and other calcineurin inhibitors. Patients applied topical pimecrolimus 1% cream on their skin lesions twice daily for 16 weeks (1 g per 2% of body surface), with subsequent follow-up of 12 months. Dosage modifications were not allowed. To evaluate adherence to the treatment, patients were instructed to return all empty tubes to the hospital (as per drug accountability protocols). The primary endpoint was the overall response ratein the intention-to-treat population. PimTo-MF is registered with EudraCT, 2014-001377-14, and is complete. FINDINGS: Between March 1, 2015, and Sept 30, 2016, 39 patients were enrolled. All patients were assessable, with a median age of 51·5 years (IQR 45-62), and the population was predominantly male (24 male [62%], 15 female [38%]). Median follow-up after baseline was 5·7 years (IQR 5·7-6·2). 22 (56%) of 39 patients had an overall response (one complete response, 21 partial responses). Responses were observed across IA (14 [54%] of 26 patients) and IB (eight [73%] of 11 patients) clinical stages, but not IIA. Topical pimecrolimus was well tolerated and no patient required a dose reduction or discontinued treatment because of unacceptable drug-related toxicity. No patients were lost to follow-up or discontinued treatment. 13 (33%) of 39 patients reported adverse events; transitory mild burning or pruritus (grade 1) was the most common, seen in eight (21%) patients. In three (8%) of these patients, the burning or pruritus was considered related to treatment. No grade 4 or 5 adverse events were observed. INTERPRETATION: Pimecrolimus 1% cream seems active and safe in patients with early stage mycosis fungoides. Our findings should be taken with caution until long-term follow-up data are obtained that confirm the safety of this treatment. Further controlled clinical trials are warranted to confirm these results. FUNDING: Instituto de Salud Carlos III and the European Regional Development Fund. TRANSLATION: For the Spanish translation of the abstract see Supplementary Materials section.
Asunto(s)
Micosis Fungoide , Neoplasias Cutáneas , Adolescente , Adulto , Femenino , Humanos , Masculino , Persona de Mediana Edad , Micosis Fungoide/tratamiento farmacológico , Prurito/tratamiento farmacológico , Neoplasias Cutáneas/tratamiento farmacológico , Tacrolimus/efectos adversos , Tacrolimus/análogos & derivadosRESUMEN
BACKGROUND: Eczematous external otitis is a common chronic condition that can have a significant impact on the life of sufferers, causing constant discomfort and pruritus, and leading to sleep deprivation. Treatment is based on the use of topical steroids, moisturisers and occasionally antibiotics. Results, however, can be disappointing, especially over the long term. METHODS: This study compared the long-term response to pimecrolimus, administered to a group of 11 patients, against clobetasone butyrate, administered to an equivalent number of patients. Response to the treatment was assessed and statistically analysed at 3 and 12 months. CONCLUSION: Whereas the degree of improvement following the use of pimecrolimus and clobetasone butyrate was similar for the two groups at month 3, a highly statistically significant difference was documented at month 12, with a much greater and sustained improvement in the pimecrolimus group.
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Conducto Auditivo Externo , Otitis Externa , Administración Tópica , Clobetasol/análogos & derivados , Clobetasol/uso terapéutico , Humanos , Otitis Externa/tratamiento farmacológico , Tacrolimus/análogos & derivadosRESUMEN
iCasp9 suicide gene has been widely used as a promising killing strategy in various cell therapies. However, different cells show significant heterogeneity in response to apoptosis inducer, posing challenges in clinical applications of killing strategy. The cause of the heterogeneity remains elusive so far. Here, by simultaneously monitoring the dynamics of iCasp9 dimerization, Caspase3 activation, and cell fate in single cells, we found that the heterogeneity was mainly due to cell-to-cell variability in initial iCasp9 expression and XIAP/Caspase3 ratio. Moreover, multiple-round drugging cannot increase the killing efficiency. Instead, it will place selective pressure on protein levels, especially on the level of initial iCasp9, leading to drug resistance. We further show this resistance can be largely eliminated by combinatorial drugging with XIAP inhibitor at the end, but not at the beginning, of the multiple-round treatments. Our results unveil the source of cell fate heterogeneity and drug resistance in iCasp9-mediated cell death, which may enlighten better therapeutic strategies for optimized killing.
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Caspasa 9/farmacología , Muerte Celular/efectos de los fármacos , Caspasa 3/metabolismo , Caspasa 9/química , Caspasa 9/metabolismo , Supervivencia Celular/efectos de los fármacos , Relación Dosis-Respuesta a Droga , Resistencia a Medicamentos/efectos de los fármacos , Sinergismo Farmacológico , Células HEK293 , Células HeLa , Humanos , Multimerización de Proteína , Tacrolimus/análogos & derivados , Tacrolimus/farmacología , Factores de Tiempo , Proteína Inhibidora de la Apoptosis Ligada a X/antagonistas & inhibidores , Proteína Inhibidora de la Apoptosis Ligada a X/metabolismoRESUMEN
Treatment of vitiligo represents a highly therapeutic challenge in spite of the continuous development of new modalities. Combination therapies of vitiligo can help improve treatment response, and reduce recurrence potential. To compare the efficacy and adverse effects of microneedling combined with-fluorouracil, pimecrolimus, and trichloroacetic acid (TCA) in the treatment of localized, stable vitiligo. The study included 75 patients with non-segmental, stable vitiligo who were randomly assigned to three equal groups: group received a combination of microneedling and -FU, group 2 received microneedling and pimecrolimus, and group 3 received microneedling and TCA. The procedure was done every 2 weeks for a maximum of six sessions. Combined microneedling and TCA was associated with the highest + 5-fluorouracil, and lastly combined microneedling + pimecrolimus. The difference between the three groups was statistically significant in favor of the combined microneedling and TCA. Pain, erythema, post-inflammatory hyperpigmentation, infection, and scarring were variably reported adverse effects in the three groups. Combination therapy seems to be a promising modality for the treatment of vitiligo. Combined microneedling and TCA is superior to combined microneedling with either-fluorouracil or pimecrolimus.
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Ácido Tricloroacético , Vitíligo , Terapia Combinada , Fluorouracilo , Humanos , Tacrolimus/análogos & derivados , Resultado del Tratamiento , Ácido Tricloroacético/efectos adversos , Vitíligo/inducido químicamente , Vitíligo/diagnóstico , Vitíligo/terapiaRESUMEN
BACKGROUND: Facial seborrhoeic dermatitis (FSD) is a chronic inflammatory skin disorder characterized by remission and exacerbation episodes. In most cases, FSD requires long-term treatment. AIM: To compare the efficacy and safety of pimecrolimus and sertaconazole in patients with FSD. METHODS: In total, 60 patients with FSD were enrolled in this double-blind, active-controlled, randomized trial, and instructed to topically apply either pimecrolimus 1% cream (30 patients) or sertaconazole 2% cream (30 patients) twice daily for 4 weeks. Assessment of disease severity was performed using the Scoring Index (SI) at baseline, on Days 14 and 28, and at 4 weeks after treatment cessation. The levels of satisfaction from treatment and any adverse effects (AEs) were also assessed in both groups. RESULTS: Although the severity of disease reduced upon treatment in both groups, application of pimecrolimus caused a significantly better improvement than sertaconazole on Days 14 and 28 (P < 0.01 and P < 0.001, respectively). The rate of relapse was significantly lower in the pimecrolimus compared with the sertaconazole group at 4 weeks after treatment cessation (P = 0.01). The highest level of satisfaction (46.7%) was observed on Day 28 in the pimecrolimus group. Both topical treatments had acceptable safety profiles; however, pimecrolimus 1% cream was significantly (P < 0.01) less irritating than sertaconazole 2% cream. CONCLUSION: Pimecrolimus is associated with faster response and fewer AEs than sertaconazole in patients with FSD.
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Dermatitis Seborreica , Fármacos Dermatológicos , Dermatitis Seborreica/tratamiento farmacológico , Fármacos Dermatológicos/uso terapéutico , Método Doble Ciego , Emolientes/uso terapéutico , Humanos , Imidazoles , Tacrolimus/efectos adversos , Tacrolimus/análogos & derivados , Tiofenos , Resultado del TratamientoRESUMEN
Acute kidney injury (AKI) is a major cause of patient mortality and a major risk multiplier for the progression to chronic kidney disease (CKD). The mechanism of the AKI to CKD transition is complex but is likely mediated by the extent and length of the inflammatory response following the initial injury. Lymphatic vessels help to maintain tissue homeostasis through fluid, macromolecule, and immune modulation. Increased lymphatic growth, or lymphangiogenesis, often occurs during inflammation and plays a role in acute and chronic disease processes. What roles renal lymphatics and lymphangiogenesis play in AKI recovery and CKD progression remains largely unknown. To determine if the increased lymphatic density is protective in the response to kidney injury, we utilized a transgenic mouse model with inducible, kidney-specific overexpression of the lymphangiogenic protein vascular endothelial growth factor-D to expand renal lymphatics. "KidVD" mouse kidneys were injured using inducible podocyte apoptosis and proteinuria (POD-ATTAC) or bilateral ischemia reperfusion. In the acute injury phase of both models, KidVD mice demonstrated a similar loss of function measured by serum creatinine and glomerular filtration rate compared to their littermates. While the initial inflammatory response was similar, KidVD mice demonstrated a shift toward more CD4+ and fewer CD8+ T cells in the kidney. Reduced collagen deposition and improved functional recovery over time was also identified in KidVD mice. In KidVD-POD-ATTAC mice, an increased number of podocytes were counted at 28 days post-injury. These data demonstrate that increased lymphatic density prior to injury alters the injury recovery response and affords protection from CKD progression.
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Lesión Renal Aguda/metabolismo , Riñón/metabolismo , Vasos Linfáticos/metabolismo , Recuperación de la Función , Daño por Reperfusión/metabolismo , Lesión Renal Aguda/etiología , Lesión Renal Aguda/inmunología , Animales , Apoptosis , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Colágeno/metabolismo , Modelos Animales de Enfermedad , Riñón/inmunología , Linfangiogénesis/genética , Ratones , Ratones Transgénicos , Podocitos , Proteinuria/inmunología , Proteinuria/metabolismo , Daño por Reperfusión/inmunología , Tacrolimus/análogos & derivados , Tacrolimus/toxicidad , Factor D de Crecimiento Endotelial Vascular/genéticaRESUMEN
Zika virus (ZIKV) infection could lead to Guillain-Barré syndrome in adults and microcephaly in the newborns from infected pregnant women. To date, there is no specific drug for the treatment of ZIKV infection. In this study, we sought to screen inhibitors against ZIKV infection from a natural product library. A ZIKV replicon was used to screen a library containing 1680 natural compounds. We explored the antiviral mechanism of the compound candidate in vitro and in vivo infection models. Ascomycin, a macrolide from Streptomyces hygroscopicus, was identified with inhibitory effect against ZIKV in Vero cells (IC50 = 0.11 µM), hepatoma cell Huh7 (IC50 = 0.38 µM), and glioblastoma cell SNB-19 (IC50 = 0.06 µM), far below the cytotoxic concentrations. Mechanistic study revealed that Ascomycin suppressed ZIKV RNA replication step during the life cycle and the regulation of calcineurin-NFAT pathway maybe involved in this inhibitory effect, independent of innate immunity activation. Moreover, we found that Ascomycin also inhibited the infection of other Flaviviridae members, such as hepatitis C virus and dengue virus. Ascomycin reduced ZIKV load in blood by up to 3500-fold in A129 mice. Meanwhile, the infection in the mice brain was undetectable by immunohistochemistry staining. Together, these findings reveal a critical role of Ascomycin in the inhibition of ZIKV and related viruses, facilitating the development of novel antiviral agents.
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Antivirales/farmacología , Productos Biológicos/farmacología , Ensayos Analíticos de Alto Rendimiento/métodos , Tacrolimus/análogos & derivados , Replicación Viral/efectos de los fármacos , Virus Zika/efectos de los fármacos , Animales , Antivirales/uso terapéutico , Productos Biológicos/química , Productos Biológicos/aislamiento & purificación , Productos Biológicos/uso terapéutico , Chlorocebus aethiops , Descubrimiento de Drogas , Humanos , Concentración 50 Inhibidora , Ratones , Bibliotecas de Moléculas Pequeñas/farmacología , Tacrolimus/aislamiento & purificación , Tacrolimus/farmacología , Tacrolimus/uso terapéutico , Células Vero , Infección por el Virus Zika/tratamiento farmacológicoRESUMEN
Drugs are designed and validated based on physicochemical data on their interactions with target proteins. For low water-solubility drugs, however, quantitative analysis is practically impossible without accurate estimation of precipitation. Here we combined quantitative NMR with NMR titration experiments to rigorously quantify the interaction of the low water-solubility drug pimecrolimus with its target protein FKBP12. Notably, the dissociation constants estimated with and without consideration of precipitation differed by more than tenfold. Moreover, the method enabled us to quantitate the FKBP12-pimecrolimus interaction even under a crowded condition established using the protein crowder BSA. Notably, the FKBP12-pimecrolimus interaction was slightly hampered under the crowded environment, which is explained by transient association of BSA with the drug molecules. Collectively, the described method will contribute to both quantifying the binding properties of low water-solubility drugs and to elucidating the drug behavior in complex crowded solutions including living cells.
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Albúmina Sérica Bovina/química , Proteína 1A de Unión a Tacrolimus/química , Tacrolimus/análogos & derivados , Animales , Bovinos , Espectroscopía de Resonancia Magnética , Solubilidad , Tacrolimus/química , Agua/químicaRESUMEN
BACKGROUND: Human mesenchymal stem cells (hMSCs) therapy has recently been considered a promising treatment for atopic dermatitis (AD) due to their immunomodulation and tissue regeneration ability. In our previous studies, we demonstrated that hMSCs alleviate allergic inflammation in murine AD model by inhibiting the activation of mast cells and B cells. Also our phase I/IIa clinical trial showed clinical efficacy and safety of hMSCs in moderate-to-severe adult AD patients. However, hMSCs therapy against atopic dermatitis have had poor results in clinical field. Therefore, we investigated the reason behind this result. We hypothesized that drug-cell interaction could interfere with the therapeutic efficacy of stem cells, and investigated whether coadministration with pimecrolimus, one of the topical calcineurin inhibitors, could influence the therapeutic potential of human umbilical cord blood mesenchymal stem cells (hUCB-MSCs) in AD. METHODS: hUCB-MSCs were subcutaneously injected to AD-induced mice with or without pimecrolimus topical application. To examine whether pimecrolimus influenced the immunomodulatory activity of hUCB-MSCs, hUCB-MSCs were treated with pimecrolimus. RESULTS: Pimecrolimus disturbed the therapeutic effect of hUCB-MSCs when they were co-administered in murine AD model. Moreover, the inhibitory functions of hUCB-MSCs against type 2 helper T (Th2) cell differentiation and mast cell activation were also deteriorated by pimecrolimus treatment. Interestingly, we found that pimecrolimus decreased the production of PGE2, one of the most critical immunomodulatory factors in hUCB-MSCs. And we demonstrated that pimecrolimus downregulated COX2-PGE2 axis by inhibiting nuclear translocation of NFAT3. CONCLUSIONS: Coadministration of pimecrolimus with hMSCs could interfere with the therapeutic efficacy of hMSCs in atopic dermatitis, and this is the first study that figured out the interaction of hMSCs with other drugs in cell therapy of atopic dermatitis. Therefore, this study might give rise to improvement of the clinical application of hMSCs therapy and facilitate the widespread application of hMSCs in clinical field.
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Dermatitis Atópica , Células Madre Mesenquimatosas , Animales , Ciclooxigenasa 2 , Dermatitis Atópica/tratamiento farmacológico , Humanos , Ratones , Tacrolimus/análogos & derivados , Tacrolimus/farmacologíaRESUMEN
FK1706 is a novel non-immunosuppressive immunophilin ligand with neurotrophic activity and exerts its neurotrophic effect through NGF. The present study aimed to elaborate on the neurotrophic activity and the mechanism of action of FK1706 in end-to-side neurorrhaphy rats and SH-SY5Y cells. In the regenerating nerves of neurorrhaphy rats, FK1706 increased the thickness of myelin sheath and the level of nerve regeneration-related proteins. The mechanism of action of FK1706 on neurite regrowth was elucidated in vitro by incubating SH-SY5Y cells in different conditions (Control, NGF, FK1706, NGF + FK1706, NGF + FK1706 + geldanamycin). Under the conditions where NGF was used, the phosphorylation level of major proteins (Raf-1 and ERK) in the Ras/Raf/MAPK/ERK signaling pathway related to SH-SY5Y cell proliferation was significantly enhanced following the application of FK1706. The number of viable cells, cell viability and neurite length of SH-SY5Y cells was maximal when NGF and FK1706 were used simultaneously. The binding level of HSP90 and Raf-1 in FK1706 group was the highest. These results indicated that FK1706 could significantly promote nerve regeneration after neurorrhaphy. The putative mechanism of action stated that FK1706 could promote the binding of HSP90 and Raf-1, make Raf-1 continue to be activated, thereby affecting key proteins in the Ras/Raf/MAPK/ERK signaling pathway related to the neurotrophic effects of NGF to promote the proliferation and neurite regrowth of nerve cells.
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Factor de Crecimiento Nervioso/farmacología , Regeneración Nerviosa/efectos de los fármacos , Raíces Nerviosas Espinales/efectos de los fármacos , Raíces Nerviosas Espinales/cirugía , Tacrolimus/análogos & derivados , Animales , Línea Celular Tumoral , Proliferación Celular/efectos de los fármacos , Proliferación Celular/fisiología , Humanos , Masculino , Modelos Animales , Factores de Crecimiento Nervioso/farmacología , Regeneración Nerviosa/fisiología , Neuritas/efectos de los fármacos , Neuritas/fisiología , Neuronas/efectos de los fármacos , Neuronas/fisiología , Ratas , Ratas Sprague-Dawley , Raíces Nerviosas Espinales/fisiología , Tacrolimus/farmacologíaRESUMEN
BACKGROUND: Peripheral vascular diseases may induce chronic ischemia and cellular injury distal to the arterial obstruction. Cellular senescence involves proliferation arrest in response to stress, which can damage neighboring cells. Renal artery stenosis (RAS) induces stenotic-kidney dysfunction and injury, but whether these arise from cellular senescenceand their temporal pattern remain unknown. METHODS: Chronic renal ischemia was induced in transgenic INK-ATTAC and wild type C57BL/6 mice by unilateral RAS, and kidney function (in vivo micro-MRI) and tissue damage were assessed. Mouse healthy and stenotic kidneys were analyzed using unbiased single-cell RNA-sequencing. To demonstrate translational relevance, cellular senescence was studied in human stenotic kidneys. RESULTS: Using intraperitoneal AP20187 injections starting 1, 2, or 4 weeks after RAS, selective clearance of cells highly expressing p16Ink4a attenuated cellular senescence and improved stenotic-kidney function; however, starting treatment immediately after RAS induction was unsuccessful. Broader clearance of senescent cells, using the oral senolytic combination dasatinib and quercetin, in C57BL/6 RAS mice was more effective in clearing cells positive for p21 (Cdkn1a) and alleviating renal dysfunction and damage. Unbiased, single-cell RNA sequencing in freshly dissociated cells from healthy and stenotic mouse kidneys identified stenotic-kidney epithelial cells undergoing both mesenchymal transition and senescence. As in mice, injured human stenotic kidneys exhibited cellular senescence, suggesting this process is conserved. CONCLUSIONS: Maladaptive tubular cell senescence, involving upregulated p16 (Cdkn2a), p19 (Cdkn2d), and p21 (Cdkn1a) expression, is associated with renal dysfunction and injury in chronic ischemia. These findings support development of senolytic strategies to delay chronic ischemic renal injury.
Asunto(s)
Senescencia Celular/fisiología , Inhibidor p16 de la Quinasa Dependiente de Ciclina/metabolismo , Isquemia/fisiopatología , Riñón/fisiopatología , Insuficiencia Renal Crónica/fisiopatología , Quinasas p21 Activadas/metabolismo , Animales , Apoptosis/efectos de los fármacos , Caspasa 8/metabolismo , Senescencia Celular/efectos de los fármacos , Senescencia Celular/genética , Enfermedad Crónica , Inhibidor p16 de la Quinasa Dependiente de Ciclina/genética , Inhibidor p19 de las Quinasas Dependientes de la Ciclina/metabolismo , Inhibidor p21 de las Quinasas Dependientes de la Ciclina/genética , Dasatinib/farmacología , Modelos Animales de Enfermedad , Activación Enzimática/efectos de los fármacos , Células Epiteliales/fisiología , Transición Epitelial-Mesenquimal , Expresión Génica , Factor de Crecimiento Similar a EGF de Unión a Heparina/genética , Humanos , Isquemia/etiología , Riñón/irrigación sanguínea , Riñón/patología , Masculino , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Osteopontina/genética , Inhibidores de Proteínas Quinasas/farmacología , Obstrucción de la Arteria Renal/complicaciones , Insuficiencia Renal Crónica/etiología , Insuficiencia Renal Crónica/patología , Análisis de Secuencia de ARN , Análisis de la Célula Individual , Tacrolimus/análogos & derivados , Tacrolimus/farmacología , Regulación hacia Arriba , Quinasas p21 Activadas/genéticaRESUMEN
Acute kidney injury (AKI) is a life-threatening disease with no known curative or preventive therapies. Data from multiple animal models and human studies have linked dysregulation of bone morphogenetic protein (BMP) signaling to AKI. Small molecules that potentiate endogenous BMP signaling should have a beneficial effect in AKI. We performed a high-throughput phenotypic screen and identified a series of FK506 analogs that act as potent BMP potentiators by sequestering FKBP12 from BMP type I receptors. We further showed that calcineurin inhibition was not required for this activity. We identified a calcineurin-sparing FK506 analog oxtFK through late-stage functionalization and structure-guided design. OxtFK demonstrated an improved safety profile in vivo relative to FK506. OxtFK stimulated BMP signaling in vitro and in vivo and protected the kidneys in an AKI mouse model, making it a promising candidate for future development as a first-in-class therapeutic for diseases with dysregulated BMP signaling.