Edwardsiella tarda Attenuates Virulence upon Oxytetracycline Resistance.

The relationship between antibiotic resistance and bacterial virulence has not yet been fully explored. Here, we use Edwardsiella tarda as the research model to investigate the proteomic change upon oxytetracycline resistance (LTB4-ROTC). Compared to oxytetracycline-sensitive E. tarda (LTB4-S), LTB4-ROTC has 234 differentially expressed proteins, of which the abundance of 84 proteins is downregulated and 15 proteins are enriched to the Type III secretion system, Type VI secretion system, and flagellum pathways. Functional analysis confirms virulent phenotypes, including autoaggregation, biofilm formation, hemolysis, swimming, and swarming, are impaired in LTB4-ROTC. Furthermore, the in vivo bacterial challenge in both tilapia and zebrafish infection models suggests that the virulence of LTB4-ROTC is attenuated. Analysis of immune gene expression shows that LTB4-ROTC induces a stronger immune response in the spleen but a weaker response in the head kidney than that induced by LTB4-S, suggesting it's a potential vaccine candidate. Zebrafish and tilapia were challenged with a sublethal dose of LTB4-ROTC as a live vaccine followed by LTB4-S challenge. The relative percentage of survival of zebrafish is 60% and that of tilapia is 75% after vaccination. Thus, our study suggests that bacteria that acquire antibiotic resistance may attenuate virulence, which can be explored as a potential live vaccine to tackle bacterial infection in aquaculture.

Wnt5a-mediated autophagy contributes to the epithelial-mesenchymal transition of human bronchial epithelial cells during asthma.

BACKGROUND: The epithelial-mesenchymal transition (EMT) of human bronchial epithelial cells (HBECs) is essential for airway remodeling during asthma. Wnt5a has been implicated in various lung diseases, while its role in the EMT of HBECs during asthma is yet to be determined. This study sought to define whether Wnt5a initiated EMT, leading to airway remodeling through the induction of autophagy in HBECs. METHODS: Microarray analysis was used to investigate the expression change of WNT5A in asthma patients. In parallel, EMT models were induced using 16HBE cells by exposing them to house dust mites (HDM) or interleukin-4 (IL-4), and then the expression of Wnt5a was observed. Using in vitro gain- and loss-of-function approaches via Wnt5a mimic peptide FOXY5 and Wnt5a inhibitor BOX5, the alterations in the expression of the epithelial marker E-cadherin and the mesenchymal marker protein were observed. Mechanistically, the Ca2+/CaMKII signaling pathway and autophagy were evaluated. An autophagy inhibitor 3-MA was used to examine Wnt5a in the regulation of autophagy during EMT. Furthermore, we used a CaMKII inhibitor KN-93 to determine whether Wnt5a induced autophagy overactivation and EMT via the Ca2+/CaMKII signaling pathway. RESULTS: Asthma patients exhibited a significant increase in the gene expression of WNT5A compared to the healthy control. Upon HDM and IL-4 treatments, we observed that Wnt5a gene and protein expression levels were significantly increased in 16HBE cells. Interestingly, Wnt5a mimic peptide FOXY5 significantly inhibited E-cadherin and upregulated α-SMA, Collagen I, and autophagy marker proteins (Beclin1 and LC3-II). Rhodamine-phalloidin staining showed that FOXY5 resulted in a rearrangement of the cytoskeleton and an increase in the quantity of stress fibers in 16HBE cells. Importantly, blocking Wnt5a with BOX5 significantly inhibited autophagy and EMT induced by IL-4 in 16HBE cells. Mechanistically, autophagy inhibitor 3-MA and CaMKII inhibitor KN-93 reduced the EMT of 16HBE cells caused by FOXY5, as well as the increase in stress fibers, cell adhesion, and autophagy. CONCLUSION: This study illustrates a new link in the Wnt5a-Ca2+/CaMKII-autophagy axis to triggering airway remodeling. Our findings may provide novel strategies for the treatment of EMT-related diseases.

Spatial accessibility and inequality analysis of rabies-exposed patients to rabies post-exposure prophylaxis clinics in Guangzhou City, China.

BACKGROUND: The incidence of rabies exposure is high and increasing in China, leading to an urgent demand of rabies post-exposure prophylaxis (PEP) clinics for the injured. However, the spatial accessibility and inequality of rabies-exposed patients to rabies PEP clinics is less known in China. METHODS: Based on rabies exposure data, PEP clinic data, and resident travel origin-destination (OD) matrix data in Guangzhou City, China, we first described the incidence of rabies exposure in Guangzhou from 2020 to 2022. Then, the Gaussian two-step floating catchment area method (2SFCA) was used to analyze the spatial accessibility of rabies-exposed patients to rabies PEP clinics in Guangzhou, and the Gini coefficient and Moran's I statistics were utilized to evaluate the inequality and clustering of accessibility scores. RESULTS: From 2020 to 2022, a total of 524,160 cases of rabies exposure were reported in Guangzhou, and the incidence showed a significant increasing trend, with an average annual incidence of 932.0/100,000. Spatial accessibility analysis revealed that the overall spatial accessibility scores for three scenarios (threshold of driving duration [d0] = 30 min, 45 min, and 60 min) were 0.30 (95% CI: 0.07, 0.87), 0.28 (95% CI: 0.11, 0.53) and 0.28 (95% CI: 0.14, 0.44), respectively. Conghua, Huangpu, Zengcheng and Nansha districts had the higher accessibility scores, while Haizhu, Liwan, and Yuexiu districts exhibited lower spatial accessibility scores. The Gini coefficient and Moran's I statistics showed that there were certain inequality and clustering in the accessibility to rabies PEP clinics in Guangzhou. CONCLUSIONS: This study clarifies the heterogeneity of spatial accessibility to rabies PEP clinics, and provide valuable insights for resource allocation to achieve the WHO target of zero human dog-mediated rabies deaths by 2030.

Paternal p,p'-DDE exposure and pre-pubertal high-fat diet increases the susceptibility to fertility impairment and sperm Igf2 DMR2 hypo-methylation in male offspring.

The hypothesis of paternal origins of health and disease (POHaD) indicates that paternal exposure to adverse environment could alter the epigenetic modification in germ line, increasing the disease susceptibility in offspring or even in subsequent generations. p,p'-Dichlorodiphenyldichloroethylene (p,p'-DDE) is an anti-androgenic chemical and male reproductive toxicant. Gestational p,p'-DDE exposure could impair reproductive development and fertility in male offspring. However, the effect of paternal p,p'-DDE exposure on fertility in male offspring remains uncovered. From postnatal day (PND) 35 to 119, male rats (F0) were given 10 mg/body weight (b.w.) p,p'-DDE or corn oil by gavage. Male rats were then mated with the control females to generate male offspring. On PND35, the male offspring were divided into 4 groups according whether to be given the high-fat diet (HF): corn oil treatment with control diet (C-C), p,p'-DDE treatment with control diet (DDE-C), corn oil treatment with high-fat diet (C-HF) or p,p'-DDE treatment with high-fat diet (DDE-HF) for 35 days. Our results indicated that paternal p,p'-DDE exposure did not affect the male fertility of male offspring directly, but decreased sperm quality and induced testicular apoptosis after the high-fat diet treatment. Further analysis demonstrated that paternal exposure to p,p'-DDE and pre-pubertal high-fat diet decreased sperm Igf2 DMR2 methylation and gene expression in male offspring. Hence, paternal exposure to p,p'-DDE and pre-pubertal high-fat diet increases the susceptibility to male fertility impairment and sperm Igf2 DMR2 hypo-methylation in male offspring, posing a significant implication in the disease etiology.

Diterpenoids from the Aerial Parts of Isodon serra with Selective Cytotoxic Activity.

Four new diterpenoids, isodosins A-D (1-4), together with nine known compounds (5-13) were isolated and identified from the aerial parts of Isodon serra (Maxim.) Hara. The structures of the new diterpenoids were elucidated based on the analysis of HR-ESI-MS data, 1D/2D-NMR-spectroscopic data, and electronic circular dichroism (ECD) calculations. Cytotoxicities of compounds 2, 3, 5, 6, and 9 against the HepG2 and H1975 cell lines were evaluated with the MTT assay. As a result, compounds 2, 3, and 6 revealed higher levels of cytotoxicity against HepG2 cells than against H1975 cells. Moreover, compund 6 demonstrated the most efficacy in inhibiting the proliferation of HepG2 cells, with an IC50 value of 41.13 ± 3.49 µM. This effect was achieved by inducing apoptosis in a dose-dependent manner. Furthermore, the relationships between the structures and activities of these compounds are briefly discussed.

METTL14 promotes fibroblast-like synoviocytes activation via the LASP1/SRC/AKT axis in rheumatoid arthritis.

The objective of this study is to explore the specific roles of a crucial N6-methyladenosine (m6A) methyltransferase, methyltransferase-like 14 (METTL14), in fibroblast-like synoviocytes (FLSs) activation of rheumatoid arthritis (RA). RA rat model was induced by administering intraperitoneally collagen antibody alcohol. Primary fibroblast-like synoviocytes (FLSs) were isolated from joint synovium tissues in rats. shRNA transfection tools were used to downregulate METTL14 expression in vivo and vitro. The injury of joint synovium was shown by hematoxylin and eosin (HE) staining. The cell apoptosis of FLSs was determined by flow cytometry. The levels of IL-6, IL-18, and C-X-C motif chemokine ligand (CXCL)10 in serum and culture supernatants were measured by ELISA kits. The expressions of LIM and SH3 domain protein 1 (LASP1), p-SRC/SRC, and p-AKT/AKT in FLSs and joint synovium tissues were determined by Western blots. The expression of METTL14 was greatly induced in the synovium tissues of RA rats compared with normal control rats. Compared with sh-NC-treated FLSs, METTL14 knockdown significantly increased cell apoptosis, inhibited cell migration and invasion, and suppressed the production of IL-6, IL-18, and CXCL10 induced by TNF-α. METTL14 silencing suppresses the expression of LASP1 and the activation of Src/AKT axis induced by TNF-α in FLSs. METTL14 improves the mRNA stability of LASP1 through m6A modification. In contrast, these were reversed by LASP1 overexpression. Moreover, METTL14 silencing clearly alleviates FLSs activation and inflammation in a RA rat model. These results suggested that METTL14 promotes FLSs activation and related inflammatory response via the LASP1/SRC/AKT signaling pathway and identified METTL14 as a potential target for treating RA.

Functional Proteomics Analysis of Norfloxacin-Resistant Edwardsiella tarda.

Multidrug-resistant Edwardsiella tarda threatens both sustainable aquaculture and human health, but the control measure is still lacking. In this study, we adopted functional proteomics to investigate the molecular mechanism underlying norfloxacin (NOR) resistance in E. tarda. We found that E. tarda had a global proteomic shift upon acquisition of NOR resistance, featured with increased expression of siderophore biosynthesis and Fe3+-hydroxamate transport. Thus, either inhibition of siderophore biosynthesis with salicyl-AMS or treatment with another antibiotic, kitasamycin (Kit), which was uptake through Fe3+-hydroxamate transport, enhanced NOR killing of NOR-resistant E. tarda both in vivo and in vitro. Moreover, the combination of NOR, salicyl-AMS, and Kit had the highest efficacy in promoting the killing effects of NOR than any drug alone. Such synergistic effect not only confirmed in vitro and in vivo bacterial killing assays but also applicable to other clinic E. tarda isolates. Thus, our data suggest a proteomic-based approach to identify potential targets to enhance antibiotic killing and propose an alternative way to control infection of multidrug-resistant E. tarda.

Early onset is an indication of the severity of DADA2 disease.

OBJECTIVE: To find indicators of disease severity and factors of early remission in patients with deficiency of adenosine deaminase 2 (DADA2). METHODS: We enrolled six DADA2 patients from six families. Direct sequencing of adenosine deaminase 2 gene (ADA2) was performed by Sanger analysis. A literature review was conducted for articles regarding paediatric DADA2. RESULTS: We found that more organs were involved in early-onset (≤1 year of age) than in late-onset (>1 year of age) DADA2 patients had high level inflammatory responses, such as elevated ESR, SF, serum amyloid A and CRP. Disease severity was not significantly different from missense and frameshift mutation. Early administration of TNF inhibitor might result in better remission and reduce recurrence. In the literature, four articles describing 51 paediatric DADA2 patients were identified. We also found that fever, stroke, peripheral nervous system involvement, hypogammaglobulinaemia and hypertension were more frequent in early onset DADA2 patients. CONCLUSION: Early-onset DADA2 may be more severe. Early administration of TNF inhibitor can effectively reduce recurrence and quickly alleviate the disease.

Age-related differences in clinical and laboratory characteristics of childhood-onset systemic lupus erythematosus: Pre-puberal-onset SLE is prone to delayed diagnosis.

OBJECTIVE: This study aimed to analyze age-specific characteristics of childhood-onset systemic lupus erythematosus (cSLE) at a health center in China. METHODS: The children with SLE were grouped based on age at disease-onset: pre-pubertal (≤7 years), peri-pubertal (8-13 years), and adolescence (14-18 years). The retrospective study included patients with cSLE diagnosed at the Beijing Children's Hospital between 2013 and 2021. RESULTS: A total of 675 females and 178 males were eligible for inclusion in this study. Among them, 160 patients were diagnosed during pre-puberty, 635 during peri-puberty, and 58 during adolescence. The female-to-male ratio of pre-pubertal, peri-pubertal, and adolescent diagnosis was 3.5: 1, 3.6: 1, and 7.28:1, respectively. The median time from onset to diagnosis during the pre-puberal period was 3.0 (IQR 1.0-24.0 months), which was longer than that during the peri-puberal period (1.4; IQR 0.7-4) months and adolescence (1.0; IQR 0.4-2) months (p = <.0001). The proportion of LN in patients diagnosed during the peri-puberal period (304, 46.6%) and during adolescence (27, 47.9%) was higher than that of patients diagnosed during the pre-puberal period (59, 36.9%) (p = .044). 46 (28.8%), 233 (36.7%), and 32 (55.2%) of children diagnosed during the pre-pubertal period, peri-pubertal period, and adolescence, respectively, suffered from leukopenia. CONCLUSION: The proportion of renal involvement and leukopenia in the pre-pubertal group was lower than that of the pubertal group and adolescent group. More importantly, the younger the age of the patient, the more likely the diagnosis to be delayed.

A single administration of FGF2 after renal ischemia-reperfusion injury alleviates post-injury interstitial fibrosis.

BACKGROUND: Despite lack of clinical therapy in acute kidney injury (AKI) or its progression to chronic kidney disease (CKD), administration of growth factors shows great potential in the treatment of renal repair and further fibrosis. At an early phase of AKI, administration of exogenous fibroblast growth factor 2 (FGF2) protects against renal injury by inhibition of mitochondrial damage and inflammatory response. Here, we investigated whether this treatment attenuates the long-term renal interstitial fibrosis induced by ischemia-reperfusion (I/R) injury. METHODS: Unilateral renal I/R with contralateral nephrectomy was utilized as an in vivo model for AKI and subsequent CKD. Rats were randomly divided into four groups: Sham-operation group, I/R group, I/R-FGF2 group and FGF2-3D group. These groups were monitored for up to 2 months. Serum creatinine, inflammatory response and renal histopathology changes were detected to evaluate the role of FGF2 in AKI and followed renal interstitial fibrosis. Moreover, the expression of vimentin, α-SMA, CD31 and CD34 were examined. RESULTS: Two months after I/R injury, the severity of renal interstitial fibrosis was significantly attenuated in both of I/R-FGF2 group and FGF2-3D group, compared with the I/R group. The protective effects of FGF2 administration were associated with the reduction of high-mobility group box 1 (HMGB1)-mediated inflammatory response, the inhibition of transforming growth factor beta (TGF-ß1)/Smads signaling-induced epithelial-mesenchymal transition and the maintenance of peritubular capillary structure. CONCLUSIONS: A single dose of exogenous FGF2 administration 1 h or 3 days after reperfusion inhibited renal fibrogenesis and thus blocked the transition of AKI to CKD. Our findings provided novel insight into the role of FGF signaling in AKI-to-CKD progression and underscored the potential of FGF-based therapy for this devastating disease.

Clinical characteristics of 1020 childhood-onset systemic lupus erythematosus: data from a health centre in China.

OBJECTIVES: Childhood-onset systemic lupus erythematosus (cSLE) is a multisystem autoimmune disease characterised and presents partially differently from adults. A large cSLE cohort study is lacking in China. The present study aimed to determine the clinical characteristics in a large population of patients with cSLE, and compare with adult-onset SLE (aSLE) in an SLE cohort of China. METHODS: The retrospective study included patients with cSLE diagnosed at the Beijing Children's hospital between July 2006 and October 2020. All patients met at least 4 of ACR classification criteria for SLE. In addition, data including demographic, clinical and serologic data were collected. Our data were compared with other cSLE cohorts and Chinese aSLE cohorts. RESULTS: A total of 1020 patients were included in this study, comprising 808 female and 212 male patients (female to male ratio, 3.8:1). The mean age at diagnosis of lupus was 11.1 years (range 1.0-17.2). It took on average 6 months (range 0.1-132) from first symptoms to cSLE diagnosis and over 12 months in 12% of patients. The most common primary manifestations at onset were rash (37.2%), fever (33.4%), nephropathy (14.2%) and arthritis (13.6%). The most common clinical manifestations were rash (67.9%) and fever (57.5%). 59.4% of patients had haematological involvement, 46.0% had lupus nephritis, 33.2% had arthritis. cSLE was more active and associated with more inflammation than aSLE patients. CONCLUSIONS: This study is a large single-centre study on cSLE from China and clarifies the clinical phenotype and autoantibody spectrum of cSLE. The clinical manifestations and autoantibody spectrum of cSLE are diverse, with regional and populational differences.

Parthenolide inhibits proliferation of cells infected with Kaposi's sarcoma-associated herpesvirus by suppression of the NF-κB signaling pathway.

The disease caused by Kaposi's sarcoma-associated herpesvirus (KSHV) is one of the major causes of death of individuals with acquired immunodeficiency syndrome (AIDS). Development of anti-KSHV drugs is thus crucial. In this study, we investigated the effect of parthenolide (PTL) on the proliferation and NF-κB signaling pathway of KSHV-infected cells. iSLK.219 and KSHV-infected SH-SY5Y cells (SK-RG) were treated with PTL, TaqMan real-time quantitative PCR was used to determine the number of copies of the KSHV genome, and mRNA and protein expression of KSHV genes were analyzed by real-time PCR and immunocytochemistry. A cell viability test was used to measure cell proliferation, and flow cytometry was used to examine the effect of the drug on the cell cycle. Cyclin D1, CDK6, CDK4, and NF-κB-related proteins, including IKKß, P-p65, and P-IKB-α, were detected by Western blot. The results showed that PTL altered the morphology of the cells, reduced the KSHV copy number, and suppressed the production of ORF50, K8.1, and v-GPCR mRNA and the LANA, ORF50, and K8.1 proteins. It blocked the G1 phase in iSLK.219 cells and decreased the levels of cyclin D1, CDK6, and CDK4 as well as the levels of NF-κB signaling proteins, including IKKß, P-p65, and P-IKB-α. Together, these results suggest that PTL is a candidate drug that can decrease KSHV pathogenicity by suppressing cell proliferation and inhibiting the NF-κB signaling pathway in KSHV-infected cells.

Fibroblast growth factor 10 ameliorates renal ischaemia-reperfusion injury by attenuating mitochondrial damage.

Ischaemia-reperfusion (I/R) injury is one of the leading causes of acute kidney injury (AKI). Its pathologic mechanism is quite complex, involving oxidative stress, inflammatory response, autophagy, and apoptosis. Fibroblast growth factor 10 (FGF10) and 5-hydroxydecanoate (5-HD) play essential roles in kidney injury. Rats were divided into four groups: (i) sham group, sham-operated animals with an unconstructed renal artery; (ii) I/R group, kidneys were subjected to 50 min of ischaemia followed by reperfusion for 2 days; (iii) I/R + FGF10 group, animals treated with 0.5 mg/kg FGF10 (i.p.) 1 h before ischaemia; and (iv) 5-HD group, animals treated with 5 mg/kg 5-HD (i.m.) 30 min before FGF10 treatment. Renal injury, apoptosis damage, mitochondrial oxidative damage, mitochondrial membrane potential (MMP), and expression of the ATP-sensitive K+ (KATP) channel subunit Kir6.2 were evaluated. FGF10 treatment significantly alleviated I/R-induced elevation in the serum creatinine level and the number of terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling-positive tubular cells in the kidney. In addition, FGF10 dramatically ameliorated renal mitochondrial-related damage, including reducing mitochondrial-dependent apoptosis, alleviating oxidative stress, maintaining the mitochondrial membrane potential, and opening the mitochondrial KATP channels. The protective effect of FGF10 was significantly compromised by the ATP-dependent potassium channel blocker 5-HD. Our data suggest that FGF10 offers effective protection against I/R and improves animal survival by attenuating mitochondrial damage.

Chitosan oligosaccharide attenuates acute kidney injury and renal interstitial fibrosis induced by ischemia-reperfusion.

Acute kidney injury (AKI) and renal interstitial fibrosis are global clinical syndromes associated with high morbidity and mortality. Renal ischemia-reperfusion (I/R) injury, which commonly occurs during surgery, is one of the major causes of AKI. Nevertheless, an efficient therapeutic approach for AKI and the development of renal interstitial fibrosis is still lacking due to its elusive pathogenetic mechanism. Here, we showed that chitosan oligosaccharide (COS), a natural oligomer polysaccharide degraded from chitosan, significantly attenuates I/R-induced AKI and maintains glomerular filtration function by inhibiting oxidative stress, mitochondrial damage, and excessive endoplasmic reticulum stress both in vitro and in vivo. In addition, long-term administration of COS can also attenuate the proliferation of myofibroblasts, mitigate extra cellular matrix deposition, and thus inhibit the transition of AKI to chronic kidney disease through participating in metabolic and redox biological processes. Our findings provide novel insights into the protective role of COS against acute kidney injury.

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OBJECTIVES: To explore the association between assisted reproductive technology (ART) and birth weight discordance in twins (BWDT). METHODS: A retrospective analysis was conducted on twin infants born between January 2011 and December 2020 at the Third Affiliated Hospital of Guangzhou Medical University, with complete basic birth data. The impact of ART on the occurrence of BWDT was identified by the multivariate logistic regression analysis. RESULTS: A total of 3 974 pairs of twins were included, with 1 431 conceived naturally and 2 543 through ART. Neonates in the ART group had higher birth weights than those in the naturally conceived group (P<0.001). The incidence of BWDT was lower in the ART group compared to the naturally conceived group (16.17% vs 21.09%, P<0.001). The multivariate logistic regression analysis, adjusting for confounding factors such as maternal age, parity, pre-pregnancy body mass index, gestational diabetes, hypothyroidism, gestational age, and chorionic properties, showed no significant difference in the risk of BWDT between the ART and naturally conceived groups (P>0.05). CONCLUSIONS: ART is not associated with the risk of BWDT.

L-OPA1 deficiency aggravates necroptosis of alveolar epithelial cells through impairing mitochondrial function during acute lung injury in mice.

Necroptosis, a recently described form of programmed cell death, is the main way of alveolar epithelial cells (AECs) death in acute lung injury (ALI). While the mechanism of how to trigger necroptosis in AECs during ALI has been rarely evaluated. Long optic atrophy protein 1 (L-OPA1) is a crucial mitochondrial inner membrane fusion protein, and its deficiency impairs mitochondrial function. This study aimed to investigate the role of L-OPA1 deficiency-mediated mitochondrial dysfunction in AECs necroptosis. We comprehensively investigated the detailed contribution and molecular mechanism of L-OPA1 deficiency in AECs necroptosis by inhibiting or activating L-OPA1. First, our data showed that L-OPA1 expression was downregulated in the lungs and AECs under the lipopolysaccharide (LPS) challenge. Furthermore, inhibition of L-OPA1 aggravated the pathological injury, inflammatory response, and necroptosis in the lungs of LPS-induced ALI mice. In vitro, inhibition of L-OPA1 induced necroptosis of AECs, while activation of L-OPA1 alleviated necroptosis of AECs under the LPS challenge. Mechanistically, inhibition of L-OPA1 aggravated necroptosis of AECs by inducing mitochondrial fragmentation and reducing mitochondrial membrane potential. While activation of L-OPA1 had the opposite effects. In summary, these findings indicate for the first time that L-OPA1 deficiency mediates mitochondrial fragmentation, induces necroptosis of AECs, and exacerbates ALI in mice.

Clinicopathological characteristics and prognostic analysis of tumor-infiltrating lymphocytes (TILs) in ductal carcinoma in situ (DCIS) and DCIS with microinvasion (DCIS-Mi) of the breast.

OBJECTIVE: Our purpose is to evaluate the correlation of TILs with clinicopathological characteristics and disease free survival (DFS) in DCIS and DCIS-Mi breast cancer (BC) patients. METHODS: We retrospectively reviewed the data of 360 DCIS patients and 125 DCIS-Mi patients treated by a single institution from 2016 to 2019. TILs are regarded as continuous variables and are divided into low (≤ 5%), medium (5-40%) and high (≥ 40%) for statistical analysis. RESULTS: In DCIS and DCIS-Mi patients, larger tumor size, higher nuclear grade, hormone receptor (HR) negativity and human epidermal growth factor receptor 2(HER2) overexpression are all related to high TILs (P < 0.05). In addition, compared with DCIS, DCIS-Mi patients were significantly associated with high TILs (P < 0.001). Based on the different results of the subtypes, we further studied the correlation between TILs and DFS in 279 cases of HER2+ patients (204 of DCIS; 75 of DCIS-Mi). In HER2+ group, DCIS-Mi was significantly associated with HR negativity (P = 0.015) and high TILs (P = 0.002) compared with DCIS patients. In the survival analysis, we found that TILs had no effect on the DFS of DCIS (P = 0.938), DCIS-Mi (P = 0.807), and HER2+ (P = 0.379) BC patients. In the univariate and multivariate cox regression analysis, the correlation between TILs and the prognosis of DFS has not been confirmed in the three BC groups (P > 0.05). CONCLUSION: TILs have played an non-negligible role in the progress of DCIS to DCIS-Mi, especially in HER2+ BC. The predictive and prognostic value of TILs still needs further research to confirm.

Identification of 4 subgroups in juvenile dermatomyositis by principal component analysis-based cluster analysis.

OBJECTIVES: Juvenile dermatomyositis (JDM) is an autoimmune disease characterised by a great heterogeneity in its clinical manifestations. In this study, we aimed to investigate the association between different clinical subtypes, laboratory data, and myositis antibodies of JDM. METHODS: A total of 132 JDM patients were enrolled and their medical records were retrospectively reviewed and autoantibodies tested. Twenty-one variables, including clinical manifestations and laboratory findings, were selected for analysis. We selected principal component analysis (PCA) as a pre-processing method for cluster analysis to convert the 21 original variables into independent principal components. We then conducted a PCA-based cluster analysis in order to analyse the association between patient clusters and the clinical data, laboratory data, and myositis autoantibodies. RESULTS: We identified 4 distinct JDM subgroups by PCA-based cluster analysis, namely: cluster A, JDM patients with arthralgia and intense inflammation; cluster B, JDM patients with clinical manifestations of vasculitis; cluster C, hypermyopathic JDM patients; and cluster D, JDM patients with skin involvement. There were significant differences between the 4 groups in serum alkaline phosphatase levels, usage of aggressive immunosuppressive therapy, and autoantibody expression of anti-mi2, anti-MDA5, anti-Jo1, and anti-PM-Scl100. CONCLUSIONS: We conducted cluster analysis of a cohort of JDM patients and identified 4 subgroups that represented diverse characteristics in the distribution of laboratory data and myositis autoantibodies, indicating that multidimensional assessment of clinical manifestations is highly valuable and urgently needed in JDM patients. These subgroups may contribute to individualised treatments and improved JDM patient prognosis.

SAMHD1 associates with inflammation and vasculitis in paediatric-onset systemic lupus erythematosus.

OBJECTIVES: In this study, we aimed to explore the expression of the Aicardi-Goutières syndrome (AGS) mutant gene SAMHD1 in paediatric-onset systemic lupus erythematosus (pSLE), its correlations with clinical and laboratory parameters, and the relationship between its expression and the type 1 interferon (IFN) signalling pathway. METHODS: Peripheral blood from 98 pSLE patients and 44 gender and age-matched healthy individuals were examined. Gene expression levels of SAMDH1 and interferon-stimulated genes (ISGs; MxA, IRF3 and IRF7) were evaluated using real-time RT-PCR assays. RESULTS: SAMHD1 levels in pSLE patients were significantly increased compared to those in healthy donors (p<0.001). SAMHD1 was associated with serum ferritin (r=0.221, p=0.042) in pSLE patients. SAMHD1 levels were significantly increased (p<0.05) in pSLE patients with butterfly erythema, alopecia, and photosensitivity. SAMHD1 was positively correlated with MxA, IRF3 and IRF7 levels, indicating that SAMHD1 was associated with the type 1 IFN signalling pathway. CONCLUSIONS: SAMHD1 was significantly increased and correlated with MxA, IRF3 and IRF7 in pSLE patients.

COX-2/sEH Dual Inhibitor Alleviates Hepatocyte Senescence in NAFLD Mice by Restoring Autophagy through Sirt1/PI3K/AKT/mTOR.

We previously found that the disorder of soluble epoxide hydrolase (sEH)/cyclooxygenase-2 (COX-2)-mediated arachidonic acid (ARA) metabolism contributes to the pathogenesis of the non-alcoholic fatty liver disease (NAFLD) in mice. However, the exact mechanism has not been elucidated. Accumulating evidence points to the essential role of cellular senescence in NAFLD. Herein, we investigated whether restoring the balance of sEH/COX-2-mediated ARA metabolism attenuated NAFLD via hepatocyte senescence. A promised dual inhibitor of sEH and COX-2, PTUPB, was used in our study to restore the balance of sEH/COX-2-mediated ARA metabolism. In vivo, NAFLD was induced by a high-fat diet (HFD) using C57BL/6J mice. In vitro, mouse hepatocytes (AML12) and mouse hepatic astrocytes (JS1) were used to investigate the effects of PTUPB on palmitic acid (PA)-induced hepatocyte senescence and its mechanism. PTUPB alleviated liver injury, decreased collagen and lipid accumulation, restored glucose tolerance, and reduced hepatic triglyceride levels in HFD-induced NAFLD mice. Importantly, PTUPB significantly reduced the expression of liver senescence-related molecules p16, p53, and p21 in HFD mice. In vitro, the protein levels of γH2AX, p53, p21, COX-2, and sEH were increased in AML12 hepatocytes treated with PA, while Ki67 and PCNA were significantly decreased. PTUPB decreased the lipid content, the number of ß-gal positive cells, and the expression of p53, p21, and γH2AX proteins in AML12 cells. Meanwhile, PTUPB reduced the activation of hepatic astrocytes JS1 by slowing the senescence of AML12 cells in a co-culture system. It was further observed that PTUPB enhanced the ratio of autophagy-related protein LC3II/I in AML12 cells, up-regulated the expression of Fundc1 protein, reduced p62 protein, and suppressed hepatocyte senescence. In addition, PTUPB enhanced hepatocyte autophagy by inhibiting the PI3K/AKT/mTOR pathway through Sirt1, contributing to the suppression of senescence. PTUPB inhibits the PI3K/AKT/mTOR pathway through Sirt1, improves autophagy, slows down the senescence of hepatocytes, and alleviates NAFLD.