A recombinant adenovirus(rAd)for expression of Mycobacterium tuberculosis(M.tb)c-di-AMP phosphodiesterase CnpB was constructed,and its induced humoral immune response was detected.The codon-optimized gene of M.tb CnpB was cloned into the adenoviral plasmid pcADV.The recombinant plasmid pcADV-CnpB was transfected into HEK293T cells,and expression was detected with Western blot.The recombinant plasmid pcADV-CnpB and the backbone plasmid were co-transfected into HEK293T cells to obtain the recombinant adenovirus rAd-CnpB.rAd-CnpB was amplified in HEK293T cells,and the target protein expression of rAd-CnpB was detected with Western blot and immunofluorescence.Mice were immunized with rAd-CnpB intranasally,and their sera and bronchoalveolar lavage fluid(BALF)were collected.ELISA was used to detect levels of antigen-specific antibodies.Restriction enzyme digestion and sequencing indicated that the recombinant plasmid pcADV-CnpB was successfully constructed and led to protein expression in eukaryotic cells.rAd-CnpB was packaged and produced in HEK293T cells.After amplification and purification,rAd-CnpB with a titer of 5.53×1010 PFU/mL was obtained.rAd-CnpB led to CnpB expression in HEK293T cells.Intranasal immunization with rAd-CnpB increased levels of IgG and secretory IgA in BALF and led to high levels of IgG in sera.rAd-CnpB,the recombinant adenovirus for expression of c-di-AMP phosphodiesterase CnpB was successfully constructed,and was found to induce antigen-specific humoral and mucosal immune responses through mucosal immunization.Thus,rAd-CnpB may be used in further research on new TB vaccine strategies.

Objective:To explore the clinicopathological characteristics and prognosis of fumarate hydratase-deficient uterine leiomyoma (FH-dUL).Methods:Clinical data and follow-up information for 117 patients with FH-dUL diagnosed through surgical pathology and immunohistochemistry in the First Affiliated Hospital of Nanjing Medical University from January 2020 to December 2024, were collected. A control group of 130 patients with common uterine leiomyomas was also included. The differences between the two groups in clinical, imaging, and pathological characteristics were compared. Additionally, recurrence rates, fertility outcomes for FH-dUL patients, and the incidence of renal cancer in FH germline mutation carriers were monitored.Results:(1) Comparison of clinicopathological characteristics: the median age of 117 FH-dUL patients was 35 years, and the median age at first diagnosis of uterine leiomyomas was 29 years, both significantly younger than the control group (41 and 36 years; both P<0.01). The FH-dUL group showed significantly higher incidences of uterine myomectomy, multiple leiomyomas, diffusion restriction on pelvic magnetic resonance imaging diffusion weighted imaging, and typical pathological features (candelabra-like vessels, bizarre nuclei, cytoplasmic eosinophilic globules, perinuclear halo, cellular atypia) and higher ultrasound blood flow score (all P<0.05). Of the 30 FH-dUL patients who underwent genetic testing, 9 had germline mutations, 3 had somatic mutations, and 6 had mutations of unclear origin. Among the 9 FH gene germline mutation patients, 2 had already developed renal cell carcinoma. (2) Recurrence analysis: among the 56 patients who underwent uterine myomectomy, 22 (39.3%, 22/56) experienced recurrence during follow-up, compared to 12 (21.8%, 12/55) of the 55 patients in the control group, the difference between the two groups was statistically significant ( P=0.046). Multivariate binary logistic regression analysis showed that cellular leiomyomas ( OR=9.489, 95% CI: 1.740-51.755; P=0.009) and multiple uterine leiomyomas ( OR=10.709, 95% CI: 1.354-84.683; P=0.025) were significant risk factors for recurrence in FH-dUL. (3) Fertility analysis: among the 66 FH-dUL patients who underwent fertility-preserving surgery, 16 had the intention to have fertility desire, only 2 (2/16) completed their fertility plans during follow-up. Conclusions:Clinicopathological features and imaging features help to differentiate FH-dUL from common type uterine fibroids, but lack specificity, and the diagnosis of FH-dUL is based on immunohistochemistry. The recurrence rate after resection of FH-dUL is high, and cellular and multiple leiomyomas are important predictors of recurrence. It is crucial to perform genetic testing, genetic counseling, drug treatment to prevent recurrence, fertility guidance, and long-term comprehensive management after surgery for FH-dUL management.

Objective:To explore the clinicopathological characteristics and prognosis of fumarate hydratase-deficient uterine leiomyoma (FH-dUL).Methods:Clinical data and follow-up information for 117 patients with FH-dUL diagnosed through surgical pathology and immunohistochemistry in the First Affiliated Hospital of Nanjing Medical University from January 2020 to December 2024, were collected. A control group of 130 patients with common uterine leiomyomas was also included. The differences between the two groups in clinical, imaging, and pathological characteristics were compared. Additionally, recurrence rates, fertility outcomes for FH-dUL patients, and the incidence of renal cancer in FH germline mutation carriers were monitored.Results:(1) Comparison of clinicopathological characteristics: the median age of 117 FH-dUL patients was 35 years, and the median age at first diagnosis of uterine leiomyomas was 29 years, both significantly younger than the control group (41 and 36 years; both P<0.01). The FH-dUL group showed significantly higher incidences of uterine myomectomy, multiple leiomyomas, diffusion restriction on pelvic magnetic resonance imaging diffusion weighted imaging, and typical pathological features (candelabra-like vessels, bizarre nuclei, cytoplasmic eosinophilic globules, perinuclear halo, cellular atypia) and higher ultrasound blood flow score (all P<0.05). Of the 30 FH-dUL patients who underwent genetic testing, 9 had germline mutations, 3 had somatic mutations, and 6 had mutations of unclear origin. Among the 9 FH gene germline mutation patients, 2 had already developed renal cell carcinoma. (2) Recurrence analysis: among the 56 patients who underwent uterine myomectomy, 22 (39.3%, 22/56) experienced recurrence during follow-up, compared to 12 (21.8%, 12/55) of the 55 patients in the control group, the difference between the two groups was statistically significant ( P=0.046). Multivariate binary logistic regression analysis showed that cellular leiomyomas ( OR=9.489, 95% CI: 1.740-51.755; P=0.009) and multiple uterine leiomyomas ( OR=10.709, 95% CI: 1.354-84.683; P=0.025) were significant risk factors for recurrence in FH-dUL. (3) Fertility analysis: among the 66 FH-dUL patients who underwent fertility-preserving surgery, 16 had the intention to have fertility desire, only 2 (2/16) completed their fertility plans during follow-up. Conclusions:Clinicopathological features and imaging features help to differentiate FH-dUL from common type uterine fibroids, but lack specificity, and the diagnosis of FH-dUL is based on immunohistochemistry. The recurrence rate after resection of FH-dUL is high, and cellular and multiple leiomyomas are important predictors of recurrence. It is crucial to perform genetic testing, genetic counseling, drug treatment to prevent recurrence, fertility guidance, and long-term comprehensive management after surgery for FH-dUL management.

A recombinant adenovirus(rAd)for expression of Mycobacterium tuberculosis(M.tb)c-di-AMP phosphodiesterase CnpB was constructed,and its induced humoral immune response was detected.The codon-optimized gene of M.tb CnpB was cloned into the adenoviral plasmid pcADV.The recombinant plasmid pcADV-CnpB was transfected into HEK293T cells,and expression was detected with Western blot.The recombinant plasmid pcADV-CnpB and the backbone plasmid were co-transfected into HEK293T cells to obtain the recombinant adenovirus rAd-CnpB.rAd-CnpB was amplified in HEK293T cells,and the target protein expression of rAd-CnpB was detected with Western blot and immunofluorescence.Mice were immunized with rAd-CnpB intranasally,and their sera and bronchoalveolar lavage fluid(BALF)were collected.ELISA was used to detect levels of antigen-specific antibodies.Restriction enzyme digestion and sequencing indicated that the recombinant plasmid pcADV-CnpB was successfully constructed and led to protein expression in eukaryotic cells.rAd-CnpB was packaged and produced in HEK293T cells.After amplification and purification,rAd-CnpB with a titer of 5.53×1010 PFU/mL was obtained.rAd-CnpB led to CnpB expression in HEK293T cells.Intranasal immunization with rAd-CnpB increased levels of IgG and secretory IgA in BALF and led to high levels of IgG in sera.rAd-CnpB,the recombinant adenovirus for expression of c-di-AMP phosphodiesterase CnpB was successfully constructed,and was found to induce antigen-specific humoral and mucosal immune responses through mucosal immunization.Thus,rAd-CnpB may be used in further research on new TB vaccine strategies.

Objective Vascular dementia(VD)is a clinical syndrome caused by various cerebrovascular diseases,including ischemic,hemorrhagic,and acute and chronic hypoxic cerebrovascular diseases,leading to impaired brain function and affecting patients'cognitive ability,daily life,and work abilities.Vascular dementia is a preventable and reversible form of dementia,second only to Alzheimer's disease as the second common cause of dementia.At present,the relevant pathogenesis of vascular dementia is not clear,and there is no clear treatment method.However,its pathogenesis may be related to neuroinflammation,oxidative stress,neuronal damage and white matter lesions.Its main risk factors include genetic factors,hypercholesterolemia,diabetes,hypertension,etc.Neuroinflammatory response plays a major role in the process of secondary brain injury caused by cerebral ischemia,and inflammatory factors lead to an inflammatory cascade reaction that exacerbates damage to the nervous system.Inhibiting the inflammatory pathway and reducing the expression of inflammatory factors can improve the symptoms of vascular dementia patients and animal models,indicating that neuroinflammation may play an important role in the pathogenesis of vascular dementia.This article explores the effects of Buyang Huanwu Decoction on inflammatory factors from the perspective of summarizing relevant literature in recent years.It mainly reviews the pharmacological effects of Buyang Huanwu Decoction on treating vascular dementia,the relationship between inflammatory factor levels and vascular dementia,and the prevention and treatment of vascular dementia by regulating inflammatory factor levels.

Objective To explore the efficacy and safety of recombinant human anti-severe acute respiratory syn-drome coronavirus 2(anti-SARS-CoV-2)monoclonal antibody injection(F61 injection)in the treatment of patients with coronavirus disease 2019(COVID-19)combined with renal damage.Methods Patients with COVID-19 and renal damage who visited the PLA General Hospital from January to February 2023 were selected.Subjects were randomly divided into two groups.Control group was treated with conventional anti-COVID-19 therapy,while trial group was treated with conventional anti-COVID-19 therapy combined with F61 injection.A 15-day follow-up was conducted after drug administration.Clinical symptoms,laboratory tests,electrocardiogram,and chest CT of pa-tients were performed to analyze the efficacy and safety of F61 injection.Results Twelve subjects(7 in trial group and 5 in control group)were included in study.Neither group had any clinical progression or death cases.The ave-rage time for negative conversion of nucleic acid of SARS-CoV-2 in control group and trial group were 3.2 days and 1.57 days(P=0.046),respectively.The scores of COVID-19 related target symptom in the trial group on the 3rd and 5th day after medication were both lower than those of the control group(both P＜0.05).According to the clinical staging and World Health Organization 10-point graded disease progression scale,both groups of subjects improved but didn't show statistical differences(P＞0.05).For safety,trial group didn't present any infusion-re-lated adverse event.Subjects in both groups demonstrated varying degrees of elevated blood glucose,elevated urine glucose,elevated urobilinogen,positive urine casts,and cardiac arrhythmia,but the differences were not statistica-lly significant(all P＞0.05).Conclusion F61 injection has initially demonstrated safety and clinical benefit in trea-ting patients with COVID-19 combined with renal damage.As the domestically produced drug,it has good clinical accessibility and may provide more options for clinical practice.

Acute kidney injury (AKI) is a clinical syndrome characterized by a rapid decline in renal function. Renal ischemia-reperfusion injury (RIRI) is one of the main causes of AKI with the underlying mechanism incompletely clarified. The liver X receptors (LXRs), including LXRα and LXRβ, are members of the nuclear receptor superfamily. It has been shown that LXRs play an important role in regulating glucose and lipid metabolism, cholesterol efflux, and inflammation. The purpose of this study was to explore the role and mechanism of LXRs in RIRI. We determined the effects of LXR activation on renal function and histological changes in a mouse RIRI model and a cellular model of hypoxia/reoxygenation (H/R). In vivo results showed that LXRs agonist GW3965 significantly inhibited the increase of serum creatinine and urea nitrogen levels induced by RIRI. Both HE and PAS staining of kidney tissues revealed that GW3965 alleviated the morphological damages caused by RIRI. Immunohistochemical staining showed that GW3965 mitigated 4-HNE and GRP78 levels induced by RIRI. Furthermore, TUNEL assay indicated that GW3965 reduced RIRI-induced renal cell apoptosis. Quantitative real-time PCR (qPCR) analysis revealed that GW3965 attenuated RIRI-induced IL-6 and IL-1β mRNA expression. Compared with wild-type group, LXRα gene deficiency had little effect on RIRI-associated renal functional decline and morphological damages. Additionally, in vitro study demonstrated that GW3965 alleviated H/R-induced decrease of HK-2 human renal proximal tubule cell viability and restored the activity of superoxide dismutase (SOD) after H/R. Western blot results showed that GW3965 mitigated the increase of 4-HNE and GRP78 protein expression levels after H/R; However, knockdown of LXRβ using the small interfering RNA (siRNA) technique reduced cell viability compared to GW3965-treated group. Taken together, the LXRs agonist GW3965 significantly alleviates RIRI in mice possibly by reducing apoptosis, oxidative stress, endoplasmic reticulum stress and inflammation. These results also preliminarily confirm that the renal protective effects of LXRs agonists are dependent on LXRβ.
Animals ; Liver X Receptors/genetics* ; Reperfusion Injury/prevention & control* ; Mice ; Benzoates/pharmacology* ; Benzylamines/pharmacology* ; Male ; Endoplasmic Reticulum Chaperone BiP ; Mice, Inbred C57BL ; Apoptosis ; Acute Kidney Injury/prevention & control* ; Kidney/pathology* ; Humans

Objective:To compare and analyze the differences in clinical and pathological features of uterine smooth muscle tumor of uncertain malignant potential (STUMP), common uterine leiomyoma (UL), and cellular uterine leiomyoma (CUL), and to identify biomarkers for predicting STUMP recurrence.Methods:A total of 72 cases of STUMP patients (STUMP group) treated at the First Affiliated Hospital of Nanjing Medical University and the First Affiliated Hospital of Soochow University were collected from June 2015 to March 2024. Additionally, 72 cases of UL and 72 cases of CUL (UL group and CUL group) in the same period were collected as controls. The clinical and pathological features of the three groups were compared, and the recurrence rates and related factors affecting STUMP recurrence were analyzed.Results:(1) Comparison of clinical and pathological features: there were statistically significant differences in age, history of myomectomy, and preoperative serum lactate dehydrogenase (LDH) levels among STUMP, UL, and CUL groups (all P<0.05). STUMP group were significantly older than UL group ( P<0.05). The proportions of STUMP group with a history of myomectomy and elevated preoperative serum LDH levels were significantly higher than those in UL and CUL groups (all P<0.05). On ultrasound, 16 cases of STUMP patients (22%, 16/72), 2 cases of UL patients (3%, 2/72), and 8 cases of CUL patients (11%, 18/72) had unclear fibroid borders, with significant differences between the three groups ( χ2=12.94, P=0.002), with STUMP group significantly higher than UL group ( P<0.05). Regarding immunohistochemistry, the proportion of p16 positivity, p53 mutations, and nuclear antigen associated with cell proliferation (Ki-67) >10% were significantly higher in STUMP group compared to UL group ( P<0.05). In terms of surgical approach, 52 cases of STUMP patients (72%, 52/72) underwent hysterectomy, compared to 27 cases of UL patients (38%, 27/72) and 38 cases of CUL patients (53%, 38/72), with a significant difference between the three groups ( χ2=17.89, P=0.001). The proportion of patients who underwent myomectomy was significantly lower in STUMP group compared to UL and CUL groups (both P<0.05). Among the 20 cases of STUMP patients who underwent myomectomy, 6 patients had a subsequent total hysterectomy after being diagnosed with STUMP. (2) Comparison of recurrence: the median follow-up time for the STUMP, UL, and CUL groups was 38, 12, and 29 months, respectively. During the follow-up period, 3 cases (6%, 3/53) in STUMP group, 4 cases (7%, 4/55) in UL group, and 8 cases (13%, 8/62) in CUL group had recurrence, with no significant differences between the three groups ( χ2=1.91, P=0.411). Among the 3 cases of STUMP patients with recurrence (in the pelvic cavity, liver, and abdominal wall), 2 cases had STUMP pathology on recurrence, and 1 case progressed to well-differentiated uterine leiomyosarcoma. (3) Related factors affecting STUMP recurrence: when comparing preoperative body mass index, serum LDH levels, and Ki-67 positivity ≥30% between recurrent and non-recurrent patients, significant differences were observed (all P<0.05). Univariate logistic regression analysis showed that Ki-67 positivity ≥30% was a significant risk factor for STUMP recurrence ( OR=24.67, 95% CI: 1.70-357.36, P=0.019). Conclusions:Factors such as age, history of myomectomy, preoperative serum LDH levels, and ultrasound findings of unclear fibroid borders are helpful for distinguishing STUMP from UL and CUL. Elevated preoperative serum LDH levels and Ki-67 positivity ≥30% have predictive value for STUMP recurrence. Active postoperative follow-up is essential, whether STUMP patients undergo myomectomy or hysterectomy.

Objective:To compare and analyze the differences in clinical and pathological features of uterine smooth muscle tumor of uncertain malignant potential (STUMP), common uterine leiomyoma (UL), and cellular uterine leiomyoma (CUL), and to identify biomarkers for predicting STUMP recurrence.Methods:A total of 72 cases of STUMP patients (STUMP group) treated at the First Affiliated Hospital of Nanjing Medical University and the First Affiliated Hospital of Soochow University were collected from June 2015 to March 2024. Additionally, 72 cases of UL and 72 cases of CUL (UL group and CUL group) in the same period were collected as controls. The clinical and pathological features of the three groups were compared, and the recurrence rates and related factors affecting STUMP recurrence were analyzed.Results:(1) Comparison of clinical and pathological features: there were statistically significant differences in age, history of myomectomy, and preoperative serum lactate dehydrogenase (LDH) levels among STUMP, UL, and CUL groups (all P<0.05). STUMP group were significantly older than UL group ( P<0.05). The proportions of STUMP group with a history of myomectomy and elevated preoperative serum LDH levels were significantly higher than those in UL and CUL groups (all P<0.05). On ultrasound, 16 cases of STUMP patients (22%, 16/72), 2 cases of UL patients (3%, 2/72), and 8 cases of CUL patients (11%, 18/72) had unclear fibroid borders, with significant differences between the three groups ( χ2=12.94, P=0.002), with STUMP group significantly higher than UL group ( P<0.05). Regarding immunohistochemistry, the proportion of p16 positivity, p53 mutations, and nuclear antigen associated with cell proliferation (Ki-67) >10% were significantly higher in STUMP group compared to UL group ( P<0.05). In terms of surgical approach, 52 cases of STUMP patients (72%, 52/72) underwent hysterectomy, compared to 27 cases of UL patients (38%, 27/72) and 38 cases of CUL patients (53%, 38/72), with a significant difference between the three groups ( χ2=17.89, P=0.001). The proportion of patients who underwent myomectomy was significantly lower in STUMP group compared to UL and CUL groups (both P<0.05). Among the 20 cases of STUMP patients who underwent myomectomy, 6 patients had a subsequent total hysterectomy after being diagnosed with STUMP. (2) Comparison of recurrence: the median follow-up time for the STUMP, UL, and CUL groups was 38, 12, and 29 months, respectively. During the follow-up period, 3 cases (6%, 3/53) in STUMP group, 4 cases (7%, 4/55) in UL group, and 8 cases (13%, 8/62) in CUL group had recurrence, with no significant differences between the three groups ( χ2=1.91, P=0.411). Among the 3 cases of STUMP patients with recurrence (in the pelvic cavity, liver, and abdominal wall), 2 cases had STUMP pathology on recurrence, and 1 case progressed to well-differentiated uterine leiomyosarcoma. (3) Related factors affecting STUMP recurrence: when comparing preoperative body mass index, serum LDH levels, and Ki-67 positivity ≥30% between recurrent and non-recurrent patients, significant differences were observed (all P<0.05). Univariate logistic regression analysis showed that Ki-67 positivity ≥30% was a significant risk factor for STUMP recurrence ( OR=24.67, 95% CI: 1.70-357.36, P=0.019). Conclusions:Factors such as age, history of myomectomy, preoperative serum LDH levels, and ultrasound findings of unclear fibroid borders are helpful for distinguishing STUMP from UL and CUL. Elevated preoperative serum LDH levels and Ki-67 positivity ≥30% have predictive value for STUMP recurrence. Active postoperative follow-up is essential, whether STUMP patients undergo myomectomy or hysterectomy.

Galectin-3 (Gal-3) belongs to the galectin family and is specific in binding β-galactoside. Through its C-terminal domain, Gal-3 binds to the galactoside group of the glycosylated insulin receptor (IR) and inhibits IR signaling pathway, which leads to the insulin resistance. Thus, Gal-3 is a potential therapeutic target for the treatment of insulin resistance and type 2 diabetes. Here we report a simple Gal-3 screening model based on the property that Gal-3 binds to the galactoside. We expressed and purified human Gal-3 in Escherichia coli (E.coli), and labeled it with fluorescein isothiocyanate (FITC) in vitro. After incubating FITC labeled Gal-3 (Gal-3-FITC) with PANC-1 cells, which express glycosylated membrane protein, PANC-1 cells started to show green fluorescent signal due to the Gal-3-FITC binding to the glycosylated membrane protein. Gal-3 inhibitor disrupts the binding of Gal-3-FITC and PANC1 cells, subsequently leads to the decrease of the fluorescent signal in PANC-1 cells. We can evaluate the inhibitory efficiency of Gal-3 inhibitors through measurement of the fluorescent signal. Further studies show this model is simple, stable, and repeatable with a Z' factor between 0.7 and 0.85. In sum, we have successfully established an in vitro high-throughput screening model for Gal-3 inhibitors.