RESUMO
Dermal fibroblasts (dFBs) resist infection by locally differentiating into adipocytes and producing cathelicidin antimicrobial peptide in response to Staphylococcus aureus (S. aureus). Here, we show that neonatal skin was enriched with adipogenic dFBs and immature dermal fat that highly expressed cathelicidin. The pool of adipogenic and antimicrobial dFBs declined after birth, leading to an age-dependent loss of dermal fat and a decrease in adipogenesis and cathelidicin production in response to infection. Transforming growth factor beta (TGF-ß), which acted on uncommitted embryonic and adult dFBs and inhibited their adipogenic and antimicrobial function, was identified as a key upstream regulator of this process. Furthermore, inhibition of the TGF-ß receptor restored the adipogenic and antimicrobial function of dFBs in culture and increased resistance of adult mice to S. aureus infection. These results provide insight into changes that occur in the skin innate immune system between the perinatal and adult periods of life.
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Envelhecimento/imunologia , Fibroblastos/fisiologia , Pele/metabolismo , Infecções Estafilocócicas/imunologia , Staphylococcus aureus/fisiologia , Gordura Subcutânea/metabolismo , Fator de Crescimento Transformador beta/metabolismo , Adipócitos/metabolismo , Adipogenia , Animais , Anti-Infecciosos/metabolismo , Peptídeos Catiônicos Antimicrobianos/metabolismo , Células Cultivadas , Embrião de Mamíferos , Humanos , Imunidade Inata , Camundongos , CatelicidinasRESUMO
One promising approach to overcome drug resistance in asthma treatments involves dual-target therapy, specifically targeting the ß2 adrenergic receptor (ß2-AR) and muscarinic-3 acetylcholine receptor (M3R). This study investigated the anti-asthma effects and dual-target mechanisms of glycyrrhizic acid, hesperidin, and platycodin D (GHP) from Zhisou San. GHP administration effectively attenuated OVA-induced inflammatory infiltration and overproduction of mucus in asthmatic mice. Additionally, GHP treatment significantly suppressed M3R and promoted ß2-AR activation, resulting in the relaxation of tracheal smooth muscle. These findings concluded that GHP mitigated asthma by targeting ß2-AR and M3R to ameliorate airway inflammation and modulate airway smooth muscle relaxation.
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Cerebral ischemia-reperfusion (I/R) incites neurologic damage through a myriad of complex pathophysiological mechanisms, most notably, inflammation and oxidative stress. In I/R injury, nicotinamide adenine dinucleotide phosphate (NADPH) oxidase (NOX) produces reactive oxygen species (ROS), which promote inflammatory and apoptotic pathways, augmenting ROS production and promoting cell death. Inhibiting ischemia-induced oxidative stress would be beneficial for reducing neuroinflammation and promoting neuronal cell survival. Studies have demonstrated that chlorpromazine and promethazine (C+P) induce neuroprotection. This study investigated how C+P minimizes oxidative stress triggered by ischemic injury. Adult male Sprague-Dawley rats were subject to middle cerebral artery occlusion (MCAO) and subsequent reperfusion. 8 mg/kg of C+P was injected into the rats when reperfusion was initiated. Neurologic damage was evaluated using infarct volumes, neurological deficit scoring, and TUNEL assays. NOX enzymatic activity, ROS production, protein expression of NOX subunits, manganese superoxide dismutase (MnSOD), and phosphorylation of PKC-δ were assessed. Neural SHSY5Y cells underwent oxygen-glucose deprivation (OGD) and subsequent reoxygenation and C+P treatment. We also evaluated ROS levels and NOX protein subunit expression, MnSOD, and p-PKC-δ/PKC-δ. Additionally, we measured PKC-δ membrane translocation and the level of interaction between NOX subunit (p47phox) and PKC-δ via coimmunoprecipitation. As hypothesized, treatment with C+P therapy decreased levels of neurologic damage. ROS production, NOX subunit expression, NOX activity, and p-PKC-δ/PKC-δ were all significantly decreased in subjects treated with C+P. C+P decreased membrane translocation of PKC-δ and lowered the level of interaction between p47phox and PKC-δ. This study suggests that C+P induces neuroprotective effects in ischemic stroke through inhibiting oxidative stress. Our findings also indicate that PKC-δ, NOX, and MnSOD are vital regulators of oxidative processes, suggesting that C+P may serve as an antioxidant.
Assuntos
Lesões Encefálicas , Isquemia Encefálica , AVC Isquêmico , Traumatismo por Reperfusão , Acidente Vascular Cerebral , Animais , Isquemia Encefálica/tratamento farmacológico , Clorpromazina/farmacologia , Clorpromazina/uso terapêutico , Masculino , NADPH Oxidases/metabolismo , Estresse Oxidativo , Prometazina/farmacologia , Prometazina/uso terapêutico , Ratos , Ratos Sprague-Dawley , Espécies Reativas de Oxigênio/metabolismo , Traumatismo por Reperfusão/tratamento farmacológico , Traumatismo por Reperfusão/metabolismo , Acidente Vascular Cerebral/tratamento farmacológico , Acidente Vascular Cerebral/metabolismo , Superóxido Dismutase/metabolismoRESUMO
A subset of dermal fibroblasts undergo rapid differentiation into adipocytes in response to infection and acutely produce the cathelicidin antimicrobial peptide gene Camp Vitamin A and other retinoids inhibit adipogenesis yet can show benefit to skin disorders, such as cystic acne, that are exacerbated by bacteria. We observed that retinoids potently increase and sustain the expression of Camp in preadipocytes undergoing adipogenesis despite inhibition of markers of adipogenesis, such as Adipoq, Fabp4, and Rstn Retinoids increase cathelicidin in both mouse and human preadipocytes, but this enhancement of antimicrobial peptide expression did not occur in keratinocytes or a sebocyte cell line. Preadipocytes undergoing adipogenesis more effectively inhibited growth of Staphylococcus aureus when exposed to retinoic acid. Whole transcriptome analysis identified hypoxia-inducible factor 1-α (HIF-1α) as a mechanism through which retinoids mediate this response. These observations uncouple the lipid accumulation element of adipogenesis from the innate immune response and uncover a mechanism, to our knowledge previously unsuspected, that may explain therapeutic benefits of retinoids in some skin disorders.
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Adipócitos/efeitos dos fármacos , Adipogenia/efeitos dos fármacos , Peptídeos Catiônicos Antimicrobianos/metabolismo , Derme/efeitos dos fármacos , Retinoides/farmacologia , Células 3T3-L1 , Adipócitos/metabolismo , Animais , Diferenciação Celular/efeitos dos fármacos , Linhagem Celular , Derme/metabolismo , Fibroblastos/efeitos dos fármacos , Fibroblastos/metabolismo , Queratinócitos/efeitos dos fármacos , Queratinócitos/metabolismo , Camundongos , Pele/efeitos dos fármacos , Pele/metabolismo , Infecções Estafilocócicas/tratamento farmacológico , Infecções Estafilocócicas/metabolismo , Staphylococcus aureus/efeitos dos fármacos , Tretinoína/farmacologia , CatelicidinasRESUMO
BACKGROUND: Remote ischemic preconditioning (RIPC) of a limb has been reported to protect against ischemic stroke. Our previous results demonstrated that the RIPC-mediated neuroprotection is associated with alterations in circulating immune cell populations. Here, we evaluated the effect of the spleen, the largest reservoir of immune cells, on RIPC-mediated neuroprotection against stroke. METHODS: Noninvasive RIPC was achieved by four repeated cycles of 5-min blood flow constriction in the hindlimbs using a tourniquet. The blood and spleens were collected before and 1 h and 3 days after preconditioning to analyze the effect of RIPC on the spleen and the correlation between splenic and peripheral lymphocytes. Moreover, spleen weight and splenic lymphocytes were compared in stroke rats with or without RIPC. Finally, splenectomy was made 1 day or 2 weeks before RIPC and 90-min middle cerebral artery occlusion (MCAO). The infarct areas and deficits were assessed. Blood was collected 1 h after RIPC and 3 days after MCAO to explore the impact of splenectomy on RIPC-induced neuroprotection and immune changes. The contralateral and ipsilateral hemispheres were collected 3 days after MCAO to detect the infiltration of immune cells after RIPC and splenectomy. RESULTS: Flow cytometry analysis demonstrated that the RIPC promptly increased the percentages of CD3+CD8+ cytotoxic T (Tc) cells in the spleen with a relatively delayed elevation in CD3+CD161+ natural killer T (NKT) and CD3-CD45RA+ B lymphocytes. The percentages of circulating lymphocytes are positively correlated with the percentages of splenic lymphocytes in normal rats. Interestingly, RIPC resulted in negative correlations between the percentages of splenic and circulating T lymphocytes, while the correlation between splenic and circulating B lymphocytes remained positive. For animals subjected to RIPC followed by MCAO, RIPC increased splenic volume with an expansion of splenic lymphocytes 3 days after MCAO. Furthermore, the removal of the spleen 1 day or 2 weeks before RIPC and MCAO reduced the protective effect of RIPC on ischemic brain injury and reversed the effects of RIPC on circulating immune cell composition. RIPC significantly reduced brain infiltration of Tc and NKT cells. Prior splenectomy showed no effect on immune cell infiltration after RIPC and stroke. CONCLUSION: These results reveal an immunomodulatory effect of the spleen, effecting mainly the spleen-derived lymphocytes, during RIPC-afforded neuroprotection against cerebral ischemia.
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Membro Posterior/fisiologia , Infarto da Artéria Cerebral Média/patologia , Infarto da Artéria Cerebral Média/terapia , Precondicionamento Isquêmico/métodos , Baço/fisiopatologia , Animais , Antígenos CD/metabolismo , Encéfalo/patologia , Modelos Animais de Doenças , Infarto da Artéria Cerebral Média/complicações , Leucócitos/metabolismo , Masculino , Doenças do Sistema Nervoso/etiologia , Doenças do Sistema Nervoso/prevenção & controle , Equilíbrio Postural , Ratos , Ratos Sprague-Dawley , Reperfusão , Baço/patologia , Esplenectomia , Fatores de TempoRESUMO
The effectiveness of the rehabilitative benefits of physical exercise appears to be contingent upon when the exercise is initiated after stroke. The present study assessed the hypothesis that very early exercise increases the extent of apoptotic cell death via increased expression of proapoptotic proteins in a rat stroke model. Adult male Sprague-Dawley rats were subjected to middle cerebral artery occlusion (MCAO) for 2 hr using an intraluminal filament and assigned to four nonexercise and three exercise groups. Exercise on a Rota-Rod was initiated for 30 min at 6 hr (considered very early), at 24 hr (early), and at 3 days (relatively late) after reperfusion. At 24 hr after exercise, apoptotic cell death was determined. At 3 and 24 hr after exercise, the expression of pro- and antiapoptotic proteins was evaluated through Western blotting. As expected, ischemic stroke significantly increased the levels of apoptotic cell death. Compared with the stroke group without exercise, apoptotic cell death was further increased (P < 0.05) at 6 hr but not at 24 hr or 3 days with exercise. This exacerbated cell injury was associated with increased expression of proapoptotic proteins (BAX and caspase-3). The expression of Bcl-2, an antiapoptotic protein, was not affected by exercise. In ischemic stroke, apoptotic cell death was enhanced by very early exercise in association with increased expression of proapoptotic proteins. These results shed light on the time-sensitive effect of exercise in poststroke rehabilitation. © 2016 Wiley Periodicals, Inc.
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Apoptose/fisiologia , Isquemia Encefálica/fisiopatologia , Isquemia Encefálica/reabilitação , Terapia por Exercício/métodos , Análise de Variância , Animais , Caspase 3/metabolismo , Fragmentação do DNA , Modelos Animais de Doenças , Regulação da Expressão Gênica/fisiologia , Masculino , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Ratos , Ratos Sprague-Dawley , Teste de Desempenho do Rota-Rod , Fatores de Tempo , Proteína X Associada a bcl-2/metabolismoRESUMO
Owing to the promising clinical efficacy and relatively simple composition, Shuang-Huang-Lian prescription is widely prescribed for the treatment of acute upper respiratory tract infection and acute bronchitis in practice. This necessitates the understanding of the bioactive compounds of the prescription and their binding mechanism to ß2 -adrenoceptor, which mediates the aforementioned ailments. In this work, a column containing immobilized ß2 -adrenoceptor was prepared using a diazonium salt reaction. The bioactive compound collected from the ß2 -adrenoceptor column was identified as chlorogenic acid by using high-performance liquid chromatography coupled with ion trap mass spectrometry. Using an injection amount dependent method, chlorogenic acid proved the binding to ß2 -adrenoceptor through two kinds of sites. The numbers of the sites were (1.42 ± 0.03) × 10-8 and (9.06 ± 0.49) × 10-8 M. The association constants were (2.72 ± 0.01) × 105 and (2.80 ± 0.01) × 104 M-1 , respectively. Molecular docking analysis of the interaction between chlorogenic acid and ß2 -adrenoceptor indicated that the binding mainly occurred on Ser169 , Ser173 , and Phe287 of ß2 -adrenoceptor. These results paved the way to screen bioactive compounds of other traditional medicines by receptor chromatography.
Assuntos
Medicamentos de Ervas Chinesas/química , Simulação de Acoplamento Molecular , Compostos Fitoquímicos/análise , Receptores Adrenérgicos/química , Cromatografia Líquida de Alta PressãoRESUMO
BACKGROUND: Plasmodium invasion of the mosquito midgut is a population bottleneck in the parasite lifecycle. Interference with molecular mechanisms by which the ookinete invades the mosquito midgut is one potential approach to developing malaria transmission-blocking strategies. Plasmodium aspartic proteases are one such class of potential targets: plasmepsin IV (known to be present in the asexual stage food vacuole) was previously shown to be involved in Plasmodium gallinaceum infection of the mosquito midgut, and plasmepsins VII and plasmepsin X (not known to be present in the asexual stage food vacuole) are upregulated in Plasmodium falciparum mosquito stages. These (and other) parasite-derived enzymes that play essential roles during ookinete midgut invasion are prime candidates for transmission-blocking vaccines. METHODS: Reverse transcriptase PCR (RT-PCR) was used to determine timing of P. falciparum plasmepsin VII (PfPM VII) and plasmepsin X (PfPM X) mRNA transcripts in parasite mosquito midgut stages. Protein expression was confirmed by western immunoblot and immunofluorescence assays (IFA) using anti-peptide monoclonal antibodies (mAbs) against immunogenic regions of PfPM VII and PfPM X. These antibodies were also used in standard membrane feeding assays (SMFA) to determine whether inhibition of these proteases would affect parasite transmission to mosquitoes. The Mann-Whitney U test was used to analyse mosquito transmission assay results. RESULTS: RT-PCR, western immunoblot and immunofluorescence assay confirmed expression of PfPM VII and PfPM X in mosquito stages. Whereas PfPM VII was expressed in zygotes and ookinetes, PfPM X was expressed in gametes, zygotes, and ookinetes. Antibodies against PfPM VII and PfPM X decreased P. falciparum invasion of the mosquito midgut when used at high concentrations, indicating that these proteases play a role in Plasmodium mosquito midgut invasion. Failure to generate genetic knockouts of these genes limited determination of the precise role of these proteases in parasite transmission but suggests that they are essential during the intraerythrocytic life cycle. CONCLUSIONS: PfPM VII and PfPM X are present in the mosquito-infective stages of P. falciparum. Standard membrane feeding assays demonstrate that antibodies against these proteins reduce the infectivity of P. falciparum for mosquitoes, suggesting their viability as transmission-blocking vaccine candidates. Further study of the role of these plasmepsins in P. falciparum biology is warranted.
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Ácido Aspártico Endopeptidases/metabolismo , Malária Falciparum/parasitologia , Plasmodium falciparum/metabolismo , Proteínas de Protozoários/metabolismo , Proteínas Recombinantes/metabolismo , Sequência de Aminoácidos , Animais , Anopheles/imunologia , Ácido Aspártico Endopeptidases/genética , Ácido Aspártico Endopeptidases/imunologia , Eritrócitos , Humanos , Estágios do Ciclo de Vida , Malária Falciparum/transmissão , Dados de Sequência Molecular , Plasmodium falciparum/enzimologia , Plasmodium falciparum/genética , Proteínas de Protozoários/genética , Proteínas de Protozoários/imunologia , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologiaRESUMO
A vaccine to prevent the transmission of malaria parasites from infected humans to mosquitoes is an important component for the elimination of malaria in the 21st century, yet it remains neglected as a priority of malaria vaccine development. The lead candidate for Plasmodium falciparum transmission-blocking vaccine development, Pfs25, is a sexual stage surface protein that has been produced for vaccine testing in a variety of heterologous expression systems. Any realistic malaria vaccine will need to optimize proper folding balanced against cost of production, yield, and potentially reactogenic contaminants. Here Chlamydomonas reinhardtii microalga-produced recombinant Pfs25 protein was formulated with four different human-compatible adjuvants (alum, Toll-like receptor 4 [TLR-4] agonist glucopyranosal lipid A [GLA] plus alum, squalene-oil-in-water emulsion, and GLA plus squalene-oil-in-water emulsion) and compared for their ability to induce malaria transmission-blocking antibodies. Alga-produced recombinant Pfs25 plus GLA plus squalene-oil-in-water adjuvant induced the highest titer and avidity in IgG antibodies, measured using alga-produced recombinant Pfs25 as the enzyme-linked immunosorbent assay (ELISA) antigen. These antibodies specifically reacted with the surface of P. falciparum macrogametes and zygotes and effectively prevented parasites from developing within the mosquito vector in standard membrane feeding assays. Alga-produced Pfs25 in combination with a human-compatible adjuvant composed of a TLR-4 agonist in a squalene-oil-in-water emulsion is an attractive new vaccine candidate that merits head-to-head comparison with other modalities of vaccine production and administration.
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Adjuvantes Imunológicos/administração & dosagem , Anticorpos Antiprotozoários/sangue , Culicidae/parasitologia , Vacinas Antimaláricas/imunologia , Plasmodium falciparum/imunologia , Proteínas de Protozoários/imunologia , Animais , Afinidade de Anticorpos , Chlamydomonas reinhardtii/genética , Chlamydomonas reinhardtii/metabolismo , Ensaio de Imunoadsorção Enzimática , Feminino , Humanos , Imunoglobulina G/sangue , Vacinas Antimaláricas/administração & dosagem , Vacinas Antimaláricas/genética , Vacinas Antimaláricas/isolamento & purificação , Camundongos Endogâmicos BALB C , Plasmodium falciparum/isolamento & purificação , Proteínas de Protozoários/genética , Proteínas de Protozoários/isolamento & purificação , Resultado do Tratamento , Vacinas de Subunidades Antigênicas/administração & dosagem , Vacinas de Subunidades Antigênicas/genética , Vacinas de Subunidades Antigênicas/imunologia , Vacinas de Subunidades Antigênicas/isolamento & purificação , Vacinas Sintéticas/administração & dosagem , Vacinas Sintéticas/genética , Vacinas Sintéticas/imunologia , Vacinas Sintéticas/isolamento & purificaçãoRESUMO
OBJECTIVE: To analyze the etiology of fever of unknown origin (FUO). METHODS: A total of 372 patients with FUO who hospitalized in Capital Medical University Affiliated Beijing Friendship Hospital were retrospectively analyzed from January 2003 to August 2013. All the patients were divided into two groups: group A (January 2003 - December 2007) and group B (January 2008-August 2013). Diagnosis rate, duration of hospitalization (days) and time to diagnosis between the two groups were artificially compared. RESULTS: Of the 372 FUO cases, 336 were positively diagnosed with a diagnosis rate of 90.3%. Infectious diseases were still the primary causes of FUO (60.2%), including 72 cases (32.1%) of tuberculosis. Connective tissue diseases accounted for 12.9% of the FUO cases, malignancies were 8.3%, and miscellaneous diseases were 8.9%. Yet thirty six patients (9.7%) could not be confirmed until they were discharged from hospital. The duration of fever in patients with malignancies was longer than that with infectious diseases [60.0 (30.0, 90.0) days vs 30.0 (20.0, 60.0) days, P = 0.003]. Time to diagnosis of connective tissue disease and malignancies was longer than infectious diseases [(12.0(7.3, 18.8) days and 11.0 (7.0, 18.0) vs 5.0 (3.0, 8.0) days, both P values = 0.000]. The duration of hospitalization in group A was longer than that of group B [17.0(12.0, 30.0) days vs 14.0(10.0, 20.0) days, P = 0.000]. The diagnosis rate and time to diagnosis of group A were similar with those of group B. The proportion of connective tissue diseases in group A was higher than group B (18.1% vs 9.2%,χ(2) = 6.201, P = 0.013) . The proportion of infectious disease, malignancies and miscellaneous diseases was not significantly different between the two groups. CONCLUSIONS: Infectious diseases are the major causes of FUO, and the most common cause is tuberculosis. Connective tissue diseases and malignancies are the second and third causes of FUO. The duration of fever and time to diagnosis are significantly different between the different origins.
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Febre de Causa Desconhecida/etiologia , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Estudos Retrospectivos , Adulto JovemRESUMO
OBJECTIVE: Dynamic changes in ischemic pathology after stroke suggested a "critical window" of enhanced neuroplasticity immediately after stroke onset. Although physical exercise has long been considered a promising strategy of stroke rehabilitation, very early physical exercise may exacerbate brain injury. Since remote ischemic conditioning (RIC) promotes neuroprotection and neuroplasticity, the present study combined RIC with sequential exercise to establish a new rehabilitation strategy for a better rehabilitative outcome. METHODS: A total of 120 adult male Sprague-Dawley rats were used and divided into five groups: (1) sham, (2) stroke, (3) stroke with exercise, (4) stroke with RIC, and (5) stroke with RIC followed by exercise. Brain damage was evaluated by infarct volume, neurological deficit, cell death, and lactate dehydrogenase (LDH) activity. Long-term functional outcomes were determined by grid walk tests, rotarod tests, beam balance tests, forelimb placing tests, and the Morris water maze. Neuroplasticity was evaluated through measurements of both mRNA and protein levels of synaptogenesis (synaptophysin [SYN], post-synaptic density protein-95 [PSD-95], and brain-derived neurotrophic factor [BDNF]) and angiogenesis (vascular endothelial growth factor [VEGF], angiopoietin-1 [Ang-1], and angiopoietin-2 [Ang-2]). Inflammasome activation was measured by concentrations of interleukin-18 (IL-18) and IL-1ß detected by enzyme-linked immunosorbent assay (ELISA) kits, mRNA expressions of NLR pyrin domain containing 3 (NLRP3), apoptosis-associated speck-like protein containing a C-terminal caspase recruitment domain (ASC), IL-18 and IL-1ß, and protein quantities of NLRP3, ASC, cleaved-caspase-1, gasdermin D-N (GSDMD-N), and IL-18 and IL-1ß. Stress granules (SGs), including GTPase-activating protein-binding protein 1 (G3BP1), T cell-restricted intracellular antigen-1 (TIA1), and DEAD-box RNA helicase 3X (DDX3X) were evaluated at mRNA and protein levels. The interactions between DDX3X with NLRP3 or G3BP1 were determined by immunofluorescence and co-immunoprecipitation. RESULTS: Early RIC decreased infarct volumes, neurological deficits, cell death, and LDH activity at post-stroke Day 3 (p < 0.05). All treatment groups showed significant improvement in functional outcomes, including sensory, motor, and cognitive functions. RIC and exercise, as compared to RIC or physical exercise alone, had improved functional outcomes after stroke (p < 0.05), as well as synaptogenesis and angiogenesis (p < 0.05). RIC significantly reduced mRNA and protein expressions of NLRP3 (p < 0.05). SGs formation peaked at 0 h after ischemia, then progressively decreased until 24 h postreperfusion, which was reversed by RIC (p < 0.05). The assembly of SGs consumed DDX3X and then inhibited NLRP3 inflammasome activation. CONCLUSIONS: RIC followed by exercise induced a better rehabilitation in ischemic rats, while early RIC alleviated ischemia-reperfusion injury via stress-granule-mediated inhibition of NLRP3 inflammasome.
Assuntos
Lesões Encefálicas , Reabilitação do Acidente Vascular Cerebral , Acidente Vascular Cerebral , Humanos , Ratos , Masculino , Animais , Inflamassomos/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Interleucina-18/metabolismo , Ratos Sprague-Dawley , DNA Helicases/metabolismo , Grânulos de Estresse , Fator A de Crescimento do Endotélio Vascular/metabolismo , Proteínas de Ligação a Poli-ADP-Ribose , RNA Helicases/metabolismo , Proteínas com Motivo de Reconhecimento de RNA , Lesões Encefálicas/patologia , Infarto , RNA MensageiroRESUMO
BACKGROUND: Inflammation and subsequent mitochondrial dysfunction and cell death worsen outcomes after revascularization in ischemic stroke. Receptor-interacting protein kinase 1 (RIPK1) activated dynamin-related protein 1 (DRP1) in a NLRPyrin domain containing 3 (NLRP3) inflammasome-dependent fashion and Hypoxia-Inducible Factor (HIF)-1α play key roles in the process. This study determined how phenothiazine drugs (chlorpromazine and promethazine (C + P)) with the hypothermic and normothermic modality impacts the RIPK1/RIPK3-DRP1 and HIF-1α pathways in providing neuroprotection. METHODS: A total of 150 adult male Sprague-Dawley rats were subjected to 2 h middle cerebral artery occlusion (MCAO) followed by 24 h reperfusion. 8 mg/kg of C + P was administered at onset of reperfusion. Infarct volumes, mRNA and protein expressions of HIF-1α, RIPK1, RIPK3, DRP-1, NLRP3-inflammation and cytochrome c-apoptosis were assessed. Apoptotic cell death, infiltration of neutrophils and macrophages, and mitochondrial function were evaluated. Interaction between RIPK1/RIPK3 and HIF-1α/NLRP3 were determined. In SH-SY5Y cells subjected to oxygen/glucose deprivation (OGD), the normothermic effect of C + P on inflammation and apoptosis were examined. RESULTS: C + P significantly reduced infarct volumes, mitochondrial dysfunction (ATP and ROS concentration, citrate synthase and ATPase activity), inflammation and apoptosis with and without induced hypothermia. Overexpression of RIPK1, RIPK3, DRP-1, NLRP3-inflammasome and cytochrome c-apoptosis were all significantly reduced by C + P at 33 °C and the RIPK1 inhibitor (Nec1s), suggesting hypothermic effect of C + P via RIPK1/RIPK3-DRP1pathway. When body temperature was maintained at 37 °C, C + P and HIF-1α inhibitor (YC-1) reduced HIF-1α expression, leading to reduction in mitochondrial dysfunction, NLRP3 inflammasome and cytochrome c-apoptosis, as well as the interaction of HIF-1α and NLRP3. These were also evidenced in vitro, indicating a normothermic effect of C + P via HIF-1α. CONCLUSION: Hypothermic and normothermic neuroprotection of C + P involve different pathways. The normothermic effect was mediated by HIF-1α, while hypothermic effect was via RIPK1/RIPK3-DRP1 signaling. This provides a theoretical basis for future precise exploration of hypothermic and normothermic neuroprotection.
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Dinaminas , Subunidade alfa do Fator 1 Induzível por Hipóxia , Inflamassomos , AVC Isquêmico , Proteína 3 que Contém Domínio de Pirina da Família NLR , Proteína Serina-Treonina Quinases de Interação com Receptores , Transdução de Sinais , Animais , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Proteína Serina-Treonina Quinases de Interação com Receptores/genética , Proteína 3 que Contém Domínio de Pirina da Família NLR/metabolismo , Proteína 3 que Contém Domínio de Pirina da Família NLR/genética , Ratos , AVC Isquêmico/metabolismo , AVC Isquêmico/patologia , AVC Isquêmico/tratamento farmacológico , Subunidade alfa do Fator 1 Induzível por Hipóxia/metabolismo , Subunidade alfa do Fator 1 Induzível por Hipóxia/genética , Masculino , Transdução de Sinais/efeitos dos fármacos , Inflamassomos/metabolismo , Dinaminas/metabolismo , Dinaminas/genética , Ratos Sprague-Dawley , Fenotiazinas/farmacologia , Inflamação/metabolismo , Inflamação/patologia , Neuroproteção , Humanos , Modelos Animais de Doenças , Hipotermia InduzidaRESUMO
Studies have indicated that reduced serum ALT levels are commonly linked to aging and are known to predict poor outcomes in many clinical conditions as potential frailty indicators. There are close connections between the brain and peripheral organs, particularly the liver. In patients with acute ischemic stroke (AIS), the interactive effects may change ALT levels, which in turn influence stroke outcomes. Whether ALT has potential neuroprotective effects or is an indicator of frailty in AIS patients remains unknown. This retrospective analysis examined 572 AIS patients in Beijing Luhe Hospital between August 2020 and June 2021. Patient demographics and laboratory results were assembled. The National Institutes of Health Stroke Scale (NIHSS) was used to analyze stroke severity. Modified Rankin Score (mRS) determined stroke outcome 3 months after AIS, with mRS≤2 indicating a favorable outcome. Based on serum ALT measurements, patients were classified into three tertiles (T1-T3). Binary logistic regression analysis evaluated the correlation between ALT tertiles and AIS outcomes. Of the patients, 66 exhibited unfavorable outcomes. The median ALT level in this group was 13 (IQR: 11-18.25), which was lower than in the favorable outcomes cohort (16; IQR: 11-22). A decline in ALT corresponded with a higher incidence of poor outcomes at 3 months (T1, 15.5 %; T2, 11.4 %; T3, 7.0 %; p = 0.03). The lowest ALT tertile (T1) was independently linked to an adverse 3-month outcome (OR 2.50 95 %CI 1.24-5.07, p = 0.038) compared to the highest tertile. ALT levels demonstrated no correlation with age (T1, 62.59 ± 12.64; T2, 64.01 ± 11.47; T3, 65.12 ± 11.27; p > 0.05). Regardless of age, lower serum ALT levels are independently associated with poorer outcomes in AIS patients. This finding suggests the potential pivotal part of the liver in AIS outcomes, highlighting the need to consider both neurological and liver functions post-stroke.
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Background: Brain inflammation plays a key role in ischemia/reperfusion (I/R) injury and is the main cause of "ineffective or futile recanalization" after successful mechanical thrombectomy (MT) in acute ischemic stroke (AIS). One of the primary sources of inflammatory cells after AIS are derived from the spleen. As an innovative and potential neuroprotective strategy after stroke, Remote Administration of Hypothermia (RAH) temporarily suppresses immune activities in the spleen, reduces the release of inflammatory cells and cytokines into blood, and thus reversibly diminishes inflammatory injury in the brain. Methods: This single-center, prospective, randomized controlled study (RCT) is proposed for AIS patients with anterior circulation large vessel occlusion (LVO). Subjects will be randomly assigned to either the control or intervention groups in a 1:1 ratio (n = 40). Participants allocated to the intervention group will receive RAH on the abdomen above the spleen prior to recanalization until 6 h after thrombectomy. All enrolled patients will receive standard stroke Guideline care. The main adverse events associated with RAH are focal cold intolerance and abdominal pain. The primary outcome will assess safety as it pertains to RAH application. The secondary outcomes include the efficacy of RAH on spleen, determined by spleen volumes, blood inflammatory factor (cells and cytokines), and on brain injury, determined by infarction volumes and poststroke functional outcomes. Discussion: This study aims to examine the safety and preliminary effectiveness of RAH over the spleen during endovascular therapy in AIS patients. The results of this study are expected to facilitate larger randomized clinical trials and hopefully prove RAH administration confers adjuvant neuroprotective properties in AIS treated with MT. Clinical trial registration: https://www.chictr.org.cn/. Identifier ChiCTR 2300077052.
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Cutibacterium acnes is a commensal bacterium on the skin that is generally well-tolerated, but different strain types have been hypothesized to contribute to the disease acne vulgaris. To understand how some strain types might contribute to skin inflammation, we generated a repository of C. acnes isolates from skin swabs of healthy subjects and subjects with acne and assessed their strain-level identity and capacity to stimulate cytokine release. Phylotype II K-type strains were more frequent on healthy and nonlesional skin of subjects with acne than those isolated from lesions. Phylotype IA-1 C-type strains were increased on lesional skin compared with those on healthy skin. The capacity to induce cytokines from cultured monocyte-derived dendritic cells was opposite to this action on sebocytes and keratinocytes and did not correlate with the strain types associated with the disease. Whole-genome sequencing revealed a linear plasmid in high-inflammatory isolates within similar strain types that had different proinflammatory responses. Single-cell RNA sequencing of mouse skin after intradermal injection showed that strains containing this plasmid induced a higher inflammatory response in dermal fibroblasts. These findings revealed that C. acnes strain type is insufficient to predict inflammation and that carriage of a plasmid could contribute to disease.
Assuntos
Acne Vulgar , Dermatite , Animais , Camundongos , Humanos , Pele/microbiologia , Acne Vulgar/microbiologia , Propionibacterium acnes/genética , Plasmídeos/genética , Inflamação , Citocinas/genéticaRESUMO
The composition of the microbial community in the intestine may influence the functions of distant organs such as the brain, lung, and skin. These microbes can promote disease or have beneficial functions, leading to the hypothesis that microbes in the gut explain the co-occurrence of intestinal and skin diseases. Here, we show that the reverse can occur, and that skin directly alters the gut microbiome. Disruption of the dermis by skin wounding or the digestion of dermal hyaluronan results in increased expression in the colon of the host defense genes Reg3 and Muc2, and skin wounding changes the composition and behavior of intestinal bacteria. Enhanced expression Reg3 and Muc2 is induced in vitro by exposure to hyaluronan released by these skin interventions. The change in the colon microbiome after skin wounding is functionally important as these bacteria penetrate the intestinal epithelium and enhance colitis from dextran sodium sulfate (DSS) as seen by the ability to rescue skin associated DSS colitis with oral antibiotics, in germ-free mice, and fecal microbiome transplantation to unwounded mice from mice with skin wounds. These observations provide direct evidence of a skin-gut axis by demonstrating that damage to the skin disrupts homeostasis in intestinal host defense and alters the gut microbiome.
Assuntos
Colite , Microbioma Gastrointestinal , Camundongos , Animais , Ácido Hialurônico/metabolismo , Mucosa Intestinal/metabolismo , Transplante de Microbiota Fecal , Sulfato de Dextrana/toxicidade , Camundongos Endogâmicos C57BL , Modelos Animais de Doenças , Colo/metabolismoRESUMO
The skin provides an essential barrier for host defense through rapid action of multiple resident and recruited cell types, but the complex communication network governing these processes is incompletely understood. To define these cell-cell interactions more clearly, we performed an unbiased network analysis of mouse skin during invasive S. aureus infection and revealed a dominant role for CXCL12+ fibroblast subsets in neutrophil communication. These subsets predominantly reside in the reticular dermis, express adipocyte lineage markers, detect IL-17 and TNFα, and promote robust neutrophil recruitment through NFKBIZ-dependent release of CXCR2 ligands and CXCL12. Targeted deletion of Il17ra in mouse fibroblasts resulted in greatly reduced neutrophil recruitment and increased infection by S. aureus. Analogous human CXCL12+ fibroblast subsets abundantly express neutrophil chemotactic factors in psoriatic skin that are subsequently decreased upon therapeutic targeting of IL-17. These findings show that CXCL12+ dermal immune acting fibroblast subsets play a critical role in cutaneous neutrophil recruitment and host defense.
Assuntos
Interleucina-17 , Staphylococcus aureus , Camundongos , Animais , Humanos , Infiltração de Neutrófilos , Pele , Fibroblastos , Quimiocina CXCL12RESUMO
Sweet's syndrome is a poorly understood inflammatory skin disease characterized by neutrophil infiltration to the dermis. Single-nucleus and bulk transcriptomics of archival clinical samples of Sweet's syndrome revealed a prominent interferon signature in Sweet's syndrome skin that was reduced in tissue from other neutrophilic dermatoses. This signature was observed in different subsets of cells, including fibroblasts that expressed interferon-induced genes. Functionally, this response was supported by analysis of cultured primary human dermal fibroblasts that were observed to highly express neutrophil chemokines in response to activation by type I interferon. Furthermore, single-molecule resolution spatial transcriptomics of skin in Sweet's syndrome identified positionally distinct immune acting fibroblasts that included a CXCL1+ subset proximal to neutrophils and a CXCL12+ subset distal to the neutrophilic infiltrate. This study defines the cellular landscape of neutrophilic dermatoses and suggests dermal immune acting fibroblasts play a role in the pathogenesis of Sweet's syndrome through recognition of type I interferons.
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Transposon-mediated transformation was used to produce Anopheles stephensi that express single-chain antibodies (scFvs) designed to target the human malaria parasite, Plasmodium falciparum. The scFvs, m1C3, m4B7, and m2A10, are derived from mouse monoclonal antibodies that inhibit either ookinete invasion of the midgut or sporozoite invasion of salivary glands. The scFvs that target the parasite surface, m4B7 and m2A10, were fused to an Anopheles gambiae antimicrobial peptide, Cecropin A. Previously-characterized Anopheles cis-acting DNA regulatory elements were included in the transgenes to coordinate scFv production with parasite development. Gene amplification and immunoblot analyses showed promoter-specific increases in transgene expression in blood-fed females. Transgenic mosquito lines expressing each of the scFv genes had significantly lower infection levels than controls when challenged with P. falciparum.
Assuntos
Anopheles/metabolismo , Organismos Geneticamente Modificados/metabolismo , Plasmodium falciparum/metabolismo , Glândulas Salivares/metabolismo , Anticorpos de Cadeia Única/biossíntese , Animais , Anopheles/genética , Anopheles/imunologia , Feminino , Regulação da Expressão Gênica/genética , Regulação da Expressão Gênica/imunologia , Especificidade de Órgãos/genética , Especificidade de Órgãos/imunologia , Organismos Geneticamente Modificados/genética , Organismos Geneticamente Modificados/imunologia , Plasmodium falciparum/genética , Plasmodium falciparum/imunologia , Regiões Promotoras Genéticas/fisiologia , Glândulas Salivares/imunologia , Glândulas Salivares/fisiologia , Anticorpos de Cadeia Única/genética , Anticorpos de Cadeia Única/imunologia , Transgenes/fisiologiaRESUMO
Pre-stroke exercise conditioning reduces neurovascular injury and improves functional outcomes after stroke. The goal of this study was to explore if post-stroke exercise conditioning (PostE) reduced brain injury and whether it was associated with the regulation of gluconeogenesis. Adult rats received 2 h of middle cerebral artery (MCA) occlusion, followed by 24 h of reperfusion. Treadmill activity was then initiated 24 h after reperfusion for PostE. The severity of the brain damage was determined by infarct volume, apoptotic cell death, and neurological deficit at one and three days after reperfusion. We measured gluconeogenesis including oxaloacetate (OAA), phosphoenolpyruvate (PEP), pyruvic acid, lactate, ROS, and glucose via ELISA, as well as the location and expression of the key enzyme phosphoenolpyruvate carboxykinase (PCK)-1/2 via immunofluorescence. We also determined upstream pathways including forkhead transcription factor (FoxO1), p-FoxO1, 3-kinase (PI3K)/Akt, and p-PI3K/Akt via Western blot. Additionally, the cytoplasmic expression of p-FoxO1 was detected by immunofluorescence. Compared to non-exercise control, PostE (*p < .05) decreased brain infarct volumes, neurological deficits, and cell death at one and three days. PostE groups (*p < .05) saw increases in OAA and decreases in PEP, pyruvic acid, lactate, ROS, glucose levels, and tissue PCKs expression on both days. PCK-1/2 expressions were also significantly (*p < .05) suppressed by the exercise setting. Additionally, phosphorylated PI3K, AKT, and FoxO1 protein expression were significantly induced by PostE at one and three days (*p < .05). In this study, PostE reduced brain injury after stroke, in association with activated PI3K/AKT/FoxO1 signaling, and inhibited gluconeogenesis. These results suggest the involvement of FoxO1 regulation of gluconeogenesis underlying post-stroke neuroprotection.