Curcumin Induces Cycle Arrest of Colon Cancer HCT116 Cells via JAK1/STAT1/p21 Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the effect of curcumin on the cycle arrest of human colon cancer HCT116 cells and decipher the possible molecular mechanism. MethodThe methyl thiazolyl tetrazolium （MTT） method was employed to examine the effects of curcumin （0， 12.5， 25， 50， 75， 100 μmol·L-1） and 5-fluorouracil （5-FU， 600 μmol·L-1） on the proliferation of HCT116 cells at different time points （24， 48， 72 h）. Flow cytometry was employed to examine the cycle of HCT116 cells treated with curcumin （0， 25， 50， 75 μmol·L-1） and 5-FU. Western blot was employed to determine the expression of proteins in the Janus kinase 1 （JAK1）/signal transducer and activator of transcription 1 （STAT1） /cyclin-dependent kinase inhibitor 1A （p21） pathway in HCT116 cells. The binding of STAT1 to p21 promoter region was detected by chromatin immunoprecipitation （ChIP）. Small interfering RNA （siRNA） was employed to measure the role of STAT1 in regulating the expression of p21 and that of JAK1 in regulating the activation of STAT1 by Western blot and cellular immunofluorescence， respectively. ResultCompared with the blank group， the HCT-116 cells treated with curcumin and 5-FU showed decreased viability （P<0.05）， increased proportions of cells in the G0/G1 phase （P<0.05）， decreased proportions of cells in the S phase and G2/M phase （P<0.05）， down-regulated protein level of phosphorylated p21 （P<0.05）， and up-regulated protein level of p21 （P<0.05）. Compared with the curcumin group， the p21 siRNA+ curcumin group presented decreased proportion of cells in G0/G1 phase （P<0.05）. Compared with the blank group， curcumin elevated the level of phosphorylated STAT1 （p-STAT1） （P<0.05）. Compared with the curcumin group， the curcumin + STAT1 siRNA group showcased up-regulated protein level of p21 in HCT116 cells （P<0.05）. The mechanism study showed that curcumin treatment enhanced the enrichment of STAT1 in the p21 promoter region （P<0.05） compared with the blank group. Compared with the blank group， curcumin up-regulated the level of phosphorylated JAK1 （p-JAK1） （P <0.05）. Compared with the curcumin group， the curcumin + STAT1 siRNA group demonstrated up-regulated protein levels of p-STAT1 and p21 in HCT116 cells （P<0.05）. ConclusionCurcumin may induce the cycle arrest of human colon cancer HCT116 cells by activating the JAK1/STAT1/p21 signaling pathway.

Mechanism of Dihuangyin in Treatment of Mice with Atopic Dermatitis by Regulating JAK1/STAT3 Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo investigate the effect and potential mechanism of Dihuangyin on 2， 4-dinitrochlorobenzene （DNCB） -induced model mice with atopic dermatitis （AD）. MethodA mouse model with AD was established by repeatedly stimulating the back skin of mice with DNCB. After successful modeling， the mice were randomly divided into model group， Runzao group （0.78 g·kg-1）， and high， medium， and low dose （40.30， 20.15， and 10.08 g·kg-1） groups of Dihuangyin， with 12 mice in each group， and the blank group consisted of 12 mice， 72 in total. The administration groups were given the corresponding liquid by dose， and the blank group and model group were given the same dose of pure water by intragastric administration， once a day. The skin lesions and scratching times of mice were observed after continuous administration for two weeks. The back skin lesions of mice were stained with hematoxylin-eosin （HE） and toluidine blue to observe the pathology. The contents of serum immunoglobulin E （IgE）， interleukin-4 （IL-4）， interleukin-6 （IL-6）， and interferon-γ （IFN-γ） were detected by enzyme-linked immunosorbent assay （ELISA）. The mRNA expression levels of IFN-γ， IL-4， IL-6， Janus kinase 1 （JAK1）， and transcriptional activator 3 （STAT3） in skin lesion tissue were detected by Real-time fluorescence quantitative polymerase chain reaction （Real-time PCR）. The expressions of JAK1， phosphorylation（p）-JAK1， STAT3， and p-STAT3 proteins in skin lesion tissue were detected by Western blot. ResultCompared with the blank group， the back skin of the model group showed large-scale scab， dryness， erosion， hypertrophy with scratching， epidermal hyperplasia with hyperkeratosis and parakeratosis， hyperacanthosis with edema， and a large number of mast cell infiltration in the dermis， some of which were degranulated. The contents of IgE， IL-4， IL-6， and IFN-γ in the serum of mice were significantly increased （P<0.01）， and the protein expression levels of p-JAK1， STAT3， and p-STAT3 and mRNA expressions of IL-4， IL-6， IFN-γ， JAK1， and STAT3 in skin lesion tissue were significantly increased （P<0.01）. Compared with the model group， only a small amount of dryness and desquamation were observed in the back skin of mice in each administration group， and cell edema was reduced. The inflammatory infiltration was significantly reduced， and the number of mast cell infiltration was significantly decreased. The serum IgE， IL-4， IL-6， and IFN-γ of mice were decreased to varying degrees （P<0.05， P<0.01）. The protein expression levels of p-JAK1， STAT3， and p-STAT3 and mRNA expressions of IL-4， IL-6， IFN-γ， JAK1， and STAT3 in skin lesion tissue were significantly decreased， and the effect of high dose group of Dihuangyin was the best （P<0.01）. ConclusionDihuangyin can improve skin lesions and pruritus in mice with AD， and its mechanism may be related to the effective regulation of cytokines on the helper T cells （Th1）/Th2 axis by interfering with the JAK1/STAT3 signaling pathway and affecting skin barrier function.

Ameliorative Effect and Mechanism of Qingwen Baiduyin on Lipopolysaccharide-induced Acute Lung Injury / 中国实验方剂学杂志

To investigate the therapeutic effect and mechanism of Qingwen Baiduyin on acute lung injury （ALI） in mice induced by lipopolysaccharide （LPS）. MethodA total of 144 C57BL/6 mice were randomly divided into the following groups： a normal group， a model group （LPS， 5 mg·kg-1）， a dexamethasone group （5 mg·kg-1）， and low-， medium-， and high-dose Qingwen Baiduyin groups （14.105， 28.21， 56.42 g·kg-1）. The mice were treated once daily for 5 days. One hour after the final administration， the ALI model was established by intratracheal instillation of LPS， and samples were collected at 6 h and 24 h after modeling. The arterial blood gas index of mice was analyzed. The total protein content， total cell count， Evans blue dye （EBD） content， and lung tissue wet-to-dry weight ratio （W/D） of bronchoalveolar lavage fluid （BALF） were measured. Hematoxylin-eosin （HE） staining was performed to assess the pathological changes in mouse lung tissue. Western blot was used to detect the expression levels of key proteins in the Janus kinase 1/signal transducer and activator of transcription 1/interferon regulatory factor 1 （JAK1/STAT1/IRF1） signaling pathway in lung tissue. ResultCompared with the normal group， the model group showed reduced arterial oxygen pressure （pO2）， oxygen saturation （SO2）， and lung tissue W/D （P<0.05， P<0.01）， increased carbon dioxide pressure （pCO2）， total protein content， total cell count， EBD content， interferon-γ （IFN-γ）， tumor necrosis factor-α （TNF-α）， interleukin-1β （IL-1β）， chemokine CXC ligand 1 （CXCL1）， chemokine CXC ligand 2 （CXCL2）， chemokine CXC ligand 9 （CXCL9）， and chemokine CXC ligand 10 （CXCL10） content （P<0.05， P<0.01）， thickening of the alveolar walls， fusion of alveolar cavities， and infiltration of inflammatory cells in lung tissue， increased proportion of M1 macrophage polarization and lung cell apoptosis （P<0.05）， and increased protein expression levels of JAK1， phosphorylated JAK1 （p-JAK1）， inducible nitric oxide synthase （iNOS）， STAT1， phosphorylated STAT1 （p-STAT1）， IRF1， gasdermin D （GSDMD）， and mixed lineage kinase domain-like protein （MLKL） （P<0.05， P<0.01）. Compared with the model group， Qingwen Baiduyin significantly increased pO2， SO2， and lung tissue W/D （P<0.05， P<0.01）， improved the pathological changes in lung tissue， and reduced pCO2， total protein content， total cell count， EBD content， IFN-γ， TNF-α， IL-1β， CXCL1， CXCL2， CXCL9， and CXCL10 content， proportion of M1 macrophage polarization， and protein expression levels of JAK1， p-JAK1， iNOS， STAT1， p-STAT1， IRF1， GSDMD， and MLKL （P<0.05， P<0.01）. ConclusionQingwen Baiduyin can improve the lung inflammatory response and reduce lung cell apoptosis in mice with ALI by inhibiting the JAK1/STAT1/IRF1 signaling pathway， thereby exerting a lung-protective effect.

Preparation of N- 18F-fluoroethyl-tofacitinib and its application in the imaging of rheumatoid arthritis / 中华核医学与分子影像杂志

Objective:To synthesize N- 18F-fluoroethyl-tofacitinib, and explore its feasibility in the diagnosis of rheumatoid arthritis (RA). Methods:The " two-step method" was used to modify tofacitinib with 18F-fluoroethyl, and the labeling rate and radiochemical purity of the probe were measured by high performance liquid chromatography (HPLC), and the stabilities of the probe in vivo and in vitro were investigated. BALB/c mice (normal group; n=3) and collagen-induced arthritis (CIA) model mice (CIA group; n=3) were injected with N- 18F-fluoroethyl-tofacitinib and CIA model mice injected with tofacitirrib and N- 18F-fluoroethyl-tofacitinib were as blocking group ( n=3). All mice underwent microPET imaging and the percentage injection dose per gram of tissue (%ID/g) and the uptake ratio of inflamed joints to muscle (T/M) were calculated. One-way analysis of variance and the least significant difference (LSD) t test were used to analyze the data. Results:The synthesis time of N- 18F-fluoroethyl-tofacitinib was about 120 min, with the yield approximately 1%, the specific activity >13.6 GBq/μmol, and the radiochemical purity >99%. After the probe incubated with PBS, plasma or in vivo for 2 h, the radiochemical purity was still more than 95%. MicroPET imaging showed that 30 min after injection, the uptake of N- 18F-fluoroethyl-tofacitinib in the inflamed joints of CIA group was higher than that of normal group and blocking group ((10.22±1.64), (2.71±0.26) and (2.81±0.33) %ID/g; F=58.26, t values: 7.83, 7.67, P values: 0.001, 0.002). The T/M of CIA group was also higher than that of normal group and blocking group (24.73±5.77, 2.75±1.36 and 2.89±0.54; F=40.64, t values: 6.42, 6.53, P values: 0.003, 0.003). Conclusions:N- 18F-fluoroethyl-tofacitinib is successfully prepared and it is stable in vitro with good imaging performance in vivo. It may be used in clinic for the diagnosis of RA.

Molecular Mechanism of "Transmission Between Lung and Brain" of Influenza and Intervention Effect of Maxing Shigantang Based on JAK1/STAT1 Signaling Pathway / 中国实验方剂学杂志

ObjectiveTo explore the molecular mechanism of "transmission between the lung and brain" of influenza based on Janus kinase 1/signal transducer and activator of transcription 1（JAK1/STAT1） signaling pathway and further investigate the intervention effect of Maxing Shigantang （MXSGT）. MethodA total of 100 SPF BALB/c mice were randomly divided into a normal group，a model group，an oseltamivir group （21.63 mg·kg-1·d-1），an antiviral granules group（3.9 g·kg-1·d-1）， and an MXSGT group（6.05 g·kg-1·d-1）， with 20 mice in each group. The pneumonia model was induced in mice except for those in the normal group by intranasal infection of influenza A virus（IAV）. Twenty-four hours after modeling，mice were treated with corresponding drugs， while those in the normal group and the model group received the same amount of normal saline by gavage， once a day for 3 and 7 days. The pathological changes in the lung and brain were observed by hematoxylin-eosin（HE）staining. The mRNA expression of IAV nucleoprotein（NP），JAK1， and STAT1 in the lung and brain was detected by real-time quantitative polymerase chain reaction（Real-time PCR）， and the protein expression of JAK1 and STAT1 in the lung and brain was detected by Western blot. Immunohistochemical method was used to detect the expression of phosphorylated（p）-STAT1 in the lung and brain tissues， and enzyme-linked immunosorbent assay（ELISA） was used to detect the serum levels of interleukin-1β（IL-1β） and interleukin-10（IL-10）. ResultCompared with the normal group， the model group showed obvious pathological changes in the lung tissues and cerebral cortex， increased relative mRNA expression of IAV NP in the lung （P<0.01）， elevated mRNA and protein expression of JAK1 and STAT1 in the lung and brain tissues （P<0.05，P<0.01），up-regulated expression level of p-STAT1 in lung tissues and cerebral cortex （P<0.05，P<0.01）， and increased serum level of IL-1β （P<0.05）. Compared with the model group， the MXSGT group showed alleviated pathological damage to lung tissues and cerebral cortex， decreased relative mRNA expression of IAV NP in lung tissues（P<0.01），reduced mRNA and protein expression levels of JAK1 and STAT1 in lung tissues and brain tissues（P<0.05，P<0.01）， and increased serum level of IL-10（P<0.01）. ConclusionThe abnormal activation of the JAK1-STAT1 signaling pathway may be one of the molecular mechanisms of "transmission between the lung and brain" of influenza. As an effective compound prescription against the influenza virus，MXSGT can alleviate the pathological damage of brain tissues in mice infected with IAV by regulating the level of cytokines mediated by this pathway.

Efficacy and Safety of Selective Janus Kinace 1 Inhibitor Upadacitinib and Filgotinib in the Treatment of Rheumatoid Arthritis ： A Meta-analysis / 中国药房

OBJECTIVE： To evaluate the therapeutic efficacy and safety of 2 kinds of selective Janus kinase 1 （JAK-1） inhibitor Upadacitinib and Filgotinibfor in the treatment of rheumatoid arthritis， and to provide evidence-based reference for clinical treatment. METHODS： Retrieved from PubMed， Medline， Embase， the Cochrane library， CBM， CJFD， Wanfang database and VIP， RCTs about placebo （control group） versus Upadacitinib or Filgotinibfor （trial group） in the treatment of rheumatoid arthritis on the basis of methotrexate or other antirheumatic drugs were collected during the establishment of the database to Jan. 2019. Meta-analysis of therapeutic efficacy [the proportion of patients with remission rate of 20％ （ACR20）， ACR50， ACR70 according to the criteria of American Rheumatism Association， the proportion of patients with 28-joint disease activity score （DAS28）＜3.2] and safety [the incidence of adverse event （AE）， severe adverse event （SAE）， infection， severe infection， herpes zoster， liver injury] were conducted by using Rev Man 5.3 software after data extraction and quality evaluation with Cochrane system evaluator manual 5.1.0. RESULTS： A total of 8 RCTs were included， involving 2 738 patients. Meta-analysis showed that the proportion of patients with ACR20 [OR＝3.37，95％CI（2.80，4.05），P＜0.001]， ACR50 [OR＝3.78，95％CI（2.98，4.78），P＜0.001] and ACR70 [OR＝4.31，95％CI（3.05，6.09），P＜0.001]， the proportion of patients with DAS28＜3.2 [OR＝3.86，95％CI（2.98，5.00），P＜0.001]， the incidence of AE [OR＝1.33，95％CI（1.11，1.61）， P＝0.002]， the incidence of infection [OR＝1.43，95％CI（1.12，1.81），P＝0.004] in trial group were significantly higher than control group； there was no statistical significance in other indexes （P＞0.05）. CONCLUSIONS： JAK-1 inhibitors Upadacitinib and Filgotinib can improve the effect indexes of ACR20， ACR50 and ACR70 and the proportion of patients with DAS28＜3.2 of rheumatoid arthritis patients； it can not increase the incidence of SAE， severe infection， herpes zoster， liver injury， but can increase the risk of AE and infection.

Immunohistochemistry of Janus Kinase 1 (JAK1) Expression in Vitiligo

BACKGROUND: Vitiligo is a chronic autoimmune disease in which the destruction of melanocytes causes white spots on the affected skin. Janus kinase (JAK) is a family of intracellular, non-receptor tyrosine kinases that transduce cytokine-mediated signals via the JAK–signal transducer and activator of transcription pathway. The aim of the present study is to explore the possible role of JAK1 in the pathogenesis of vitiligo using immunohistochemical methods. METHODS: The current study was conducted in a sample of 39 patients who presented with vitiligo and 22 healthy individuals who were age and sex matched as a control group. We used immunohistochemistry to evaluate JAK1 status (intensity and distribution) and assess the percentage of residual melanocytes using human melanoma black 45 (HMB45). RESULTS: Intense and diffuse JAK1 expression was significantly more likely to indicate vitiliginous skin compared to normal skin (p < .001). Strong and diffuse JAK1 expression was associated with short disease duration, female sex, and lower percentage of melanocytes (detected by HMB45) (p < .05). CONCLUSIONS: JAK1 may be involved in the pathogenesis of vitiligo, as indicated by intense and diffuse expression compared to control and association with lower percentage of melanocytes detected by HMB45 immunostaining.

Expression of janus kinase 1/signal transducer and activator of transcription 1 signaling pathway in cerulean stimulated pancreatic acinar cells / 中华胰腺病杂志

Objective To investigate the relationship between the activity of Janus kinase/signal transducer and activator of transcription (JAK/STAT) signaling pathway in cerulein-induced pancreatic acinar cell line AR42J. Methods The in vivo model of AP was induced by cerulean treated pancreatic acinar cell line AR42J, then RPM and AG490 were given for intervention. Western blot was used to determine theexpressions of JAK1 and phosphorylation JAK1 ( P JAK1 ) , STAT1, PSTAT1 and TNFα, IL-1β, IL-6. The expressions of IL-6, IL-1 β, and TNFα mRNA were measured by RT-PCR. Survival rate of cells was evaluated by trypan blue stain. Results The relative expressions of JAK1, P JAK1, STAT1, P STAT1 and TNF-o, IL-1β, IL 6 without cerulean treatment were 0.09 ±0.04,0.14 ±0.08,0.21 ±0.09,0.12 ±0.12,0.10 ±0.02,0.08 ± 0.03,0.02 ± 0.02. After cerulean treatment, the expressions of abovementioned protein increased in a time-dependant manner, the expressions at 24h were 0.53 ± 0.09,0.53 ± 0.13,0.56 ± 0.09,0.55 ± 0.10,0.25 ± 0.04,0.25 ±0.09,0.27 ±0.07, which were significantly higher than those in the control group (P ＜0.05). 2 4 h after RPM and AG 4 9 0 inhibition, the expressions of TNF-α, IL-1 β, IL-6 proteins significantly decreased to 0.17 ± 0.03and 0.17 ± 0.01,0.15 ± 0.05 and 0.14 ± 0.07,0.19 ± 0.04 and 0.19 ± 0.05; their expressions of mRNA significantly decreased ( P ＜ 0.05 ). The cell survival rates in RPM and AG490 treatment group were (72.4 ± 11.2) %, (69.7 ± 9.8 ) %, and in cerulein-stimulated cells (42.2 ± 12.3 ) % ( P ＜ 0.05 ).Conclusions The JAK1/STAT1 signaling pathway was involved in pancreatic inflammatory response with cerulein stimulation. Early treatment with inhibitors to the JAK1/STAT1 signaling pathway might control the inflammatory response in acute pancreatitis.

The expression and clinical significance of SHP-1 and JAK1 mRNA in patients with acute leukemia / 白血病·淋巴瘤

Objective To investigate the expression of SHP-1 and JAK1 mRNA in acute leukemia patients and their impact on disease development,and outcome of the primary chemotherapy.Methods Semi-quantitative reverse transcription polymerase chain reaction was used to measure the expression of SHP-1 and Janus kinase 1(JAK1)mRNA in 93 patients with acute leukemia(AL)and 20 healthy adults as normal 、controls(NC).Results The expression of SHP-1 mRNA in de novo AL patients was significantly lower than that in NC group(P=0.000),which was elevated when complete remission(CR)was achieved(P=0.032)and decreased after the disease relapsed (P=0.015).The expression of JAK1 mRNA in NC group was a lower than that in de novo AL group, but with no statistical significance(P=o.051).While there was statistical significance between NC group and relapsed AL group(P=0.047).The complete remission(CR)rate of the primary chemotherapy in SHP-1 positive group Was 88.9%,but 60.38%in negative group,and there was a statistical significance between them(P=0.018).There Was a negative correlation between the expression level of SHP-1 and JAKI mRNA (P=0.048).Conclusion The expression of SHP-1 mRNA Was significantly decreased or absent in the specimens of acute leukemia patients,and the positive expression of SHP-1 mRNA may be proposed as a factor of preferable therapeutic efficacy in de novo AL and a marker for the progress of the disease.The abundance of JAK1 mRNA was possibly elevated in patients with acute leukemia.