Hepatocellular carcinoma(HCC)is one of the most common malignant tumors globally.Programmed death protein-1(PD-1)/programmed death protein ligand-1(PD-L1)inhibitors promote the reactivation of anti-tumor immune response by bloc-king the negative modulatory signaling pathway of T cells'activa-tion and inhibiting the immune escape pathway of tumor cells.PD-1/PD-L1 inhibitors become a novel therapeutic strategy to treat HCC.However,long-term clinical outcomes show that HCC patients treated with anti-PD-1/PD-L1 monotherapy still have high rates of recurrence and metastasis.Combination immuno-therapy is a novel therapeutic strategy to treat advanced HCC pa-tients,among which PD-1/PD-L1 inhibitors in combination with anti-vascular endothelial growth factor(VEGF)agents have showed promising efficacy and better safety.PD-1/PD-L1 inhib-itors plus anti-VEGF agents combined therapy inhibit the growth of hepatoma cells by participating in the cancer immunity cycle pathway.This review focuses on the research progress of PD-1/PD-L1 inhibitors,anti-VEGF agents and their combined therapy in the clinical treatment of HCC.

Objective To investigate the effectsof adipocyte-specific Structural maintenance of chromosomes 5(Smc5)knockouton glucose and lipid metabolism in mice.Methods Adipocyte-specific Smc5 knockout mice(AKO mice)were constructed based onclustered regularly interspaced short palindromic repeats/CRISPR-associated nuclease 9(CRISPR/Cas9)systems,and Smc5flox/flox mice(Flox mice)were used as controls.The Smc5 knockout efficiency of adipose tissue in mice was verified by qRT-PCR and Western blot.The body fat content was detected by dual energy X-ray absorptiometry(DEXA).The morphology of adipose tissue was observed by hematoxylin-eosin staining and the area distribution of adipocytes was calculated.TG,HDL-C,LDL-C,free fatty acid(FFA),intraperitoneal glucose tolerance test(IPGTT)and intraperitoneal insulin tolerance test(IPITT)were compared.Results The AKO mouse model of fat specific knockout of Smc5 gene was successfully constructed.Smc5 mRNA in groin fat(iWAT),epididymal fat(eWAT)and brown adipocyte(BAT)of AKO mice,body weight after 15 weeks,organ weight of iWAT at 31 weeks,organ weight of eWAT,organ weight of BAT,body weight,fat mass,fat mass/weight,LDL-C,and percentage of 15-minute blood glucose in IPITT were lower than those of Flox mice(P<0.05 or P<0.01).Conclusions Adipocyte-specific Smc5 gene knockout improves glucose and lipid metabolism by affecting adipose tissue production.

Objective To investigate the changes of macular microcirculation and aqueous humor cytokine expression in patients with diabetic macular edema(DME)after anti-vascular endothelial growth factor(VEGF)treatment,and analyze the relationship with efficacy.Methods A total of 62 patients(91 eyes)with DME who were treated in the First Hospital of Nanchang from October 2021 to August 2023 were selected and treated with intravitreal injection of conbercept.According to the reduction of central macular thickness(CMT),they were divided into efficacy significant group(CMT reduction≥100μm,59 eyes)and non-efficacy significant group(CMT reduction＜100μm or increase,32 eyes).The changes of CMT,vessel density(VD)of superficial capillary plexus(SCP),fovea avascular area(FAZ),VEGF,interleuki(IL)-6,IL-8,and IL-10 after anti-VEGF treatment were analyzed.Receiver operating characteristic(ROC)curve was used to evaluate the predictive value of each index.Results Before treatment,the levels of VEGF and IL-10 in aqueous humor in efficacy significant group were significantly higher than those in non-efficacy significant group,and the level of IL-8 was significantly lower than that in non-efficacy significant group(P＜0.05).After treatment,levels of VEGF,IL-6,IL-8 and IL-10 in aqueous humor in both groups were significantly lower than before treatment(P＜0.05).The levels of VEGF,IL-6 and IL-8 in aqueous humor in efficacy significant group were significantly lower than those in non-efficacy significant group,and the level of IL-10 was significantly higher than that in non-efficacy significant group(P＜0.05).Before and after anti-VEGF treatment,there were no significant changes in FAZ area and SCP-VD in both groups(P＞0.05).Correlation analysis showed that VEGF(r=0.571,P＜0.001)and IL-10(r=0.382,P=0.008)in aqueous humor at baseline were positively correlated with CMT reduction,IL-8 was negatively correlated with CMT reduction(r=-0.689,P＜0.001).IL-6,FAZ area and SCP-VD were not correlated with CMT reduction(P＞0.05).Cytokine levels were not correlated with FAZ area and SCP-VD(P＞0.05).ROC curve results showed that area under the curve of IL-8,VEGF and IL-10 at baseline predicting anti-VEGF efficacy were 0.825,0.813 and 0.676,respectively.Conclusion The levels of VEGF,IL-8,and IL-10 in aqueous humor at baseline in DME patients were correlated with anti-VEGF efficacy and could predict the efficacy of anti-VEGF.

Objective To investigate the clinical characteristics and prognosis of pneumococcal meningitis(PM),and drug sensitivity of Streptococcus pneumoniae(SP)isolates in Chinese children.Methods A retrospective analysis was conducted on clinical information,laboratory data,and microbiological data of 160 hospitalized children under 15 years old with PM from January 2019 to December 2020 in 33 tertiary hospitals across the country.Results Among the 160 children with PM,there were 103 males and 57 females.The age ranged from 15 days to 15 years,with 109 cases(68.1% )aged 3 months to under 3 years.SP strains were isolated from 95 cases(59.4% )in cerebrospinal fluid cultures and from 57 cases(35.6% )in blood cultures.The positive rates of SP detection by cerebrospinal fluid metagenomic next-generation sequencing and cerebrospinal fluid SP antigen testing were 40% (35/87)and 27% (21/78),respectively.Fifty-five cases(34.4% )had one or more risk factors for purulent meningitis,113 cases(70.6% )had one or more extra-cranial infectious foci,and 18 cases(11.3% )had underlying diseases.The most common clinical symptoms were fever(147 cases,91.9% ),followed by lethargy(98 cases,61.3% )and vomiting(61 cases,38.1% ).Sixty-nine cases(43.1% )experienced intracranial complications during hospitalization,with subdural effusion and/or empyema being the most common complication[43 cases(26.9% )],followed by hydrocephalus in 24 cases(15.0% ),brain abscess in 23 cases(14.4% ),and cerebral hemorrhage in 8 cases(5.0% ).Subdural effusion and/or empyema and hydrocephalus mainly occurred in children under 1 year old,with rates of 91% (39/43)and 83% (20/24),respectively.SP strains exhibited complete sensitivity to vancomycin(100% ,75/75),linezolid(100% ,56/56),and meropenem(100% ,6/6).High sensitivity rates were also observed for levofloxacin(81% ,22/27),moxifloxacin(82% ,14/17),rifampicin(96% ,25/26),and chloramphenicol(91% ,21/23).However,low sensitivity rates were found for penicillin(16% ,11/68)and clindamycin(6% ,1/17),and SP strains were completely resistant to erythromycin(100% ,31/31).The rates of discharge with cure and improvement were 22.5% (36/160)and 66.2% (106/160),respectively,while 18 cases(11.3% )had adverse outcomes.Conclusions Pediatric PM is more common in children aged 3 months to under 3 years.Intracranial complications are more frequently observed in children under 1 year old.Fever is the most common clinical manifestation of PM,and subdural effusion/emphysema and hydrocephalus are the most frequent complications.Non-culture detection methods for cerebrospinal fluid can improve pathogen detection rates.Adverse outcomes can be noted in more than 10% of PM cases.SP strains are high sensitivity to vancomycin,linezolid,meropenem,levofloxacin,moxifloxacin,rifampicin,and chloramphenicol.[Chinese Journal of Contemporary Pediatrics,2024,26(2):131-138]

Objective To analyze the clinical and serological typing characteristics of invasive Haemophilus influ-enzae(Hin)infection in children.Methods Clinical data of 34 children with invasive Hin infection admitted to Children's Hospital of Shanxi from 2015 to 2021 were analyzed retrospectively.According to clinical diagnosis,they were divided into meningitis infection group and non-meningitis infection group.General data,symptoms,signs,laboratory serological indicators,and Hin serum typing characteristics of children,as well as differences in inflammatory factor level between the two groups were analyzed.Results Among the 34 patients,22 were males and 12 were females,with a male to female ratio of 1.83∶1.Children aged ≤36 months accounted for 82.35％.The levels of procalcitonin(PCT)(23.71[4.10,77.80])ng/mL and C-reactive protein(CRP)(200.00[164.55,200.00])mg/L in children in the meningitis infection group were higher than those in the non-meningitis group(1.08[0.49,6.00]ng/mL,69.46[48.09,125.63]mg/L,respectively),with statistically significant differences(both P＜0.05).The platelet(PLT)count in the non-meningitis group([312.56±186.81]× 109/L)was higher than that in the meningitis group([183.28±165.67]× 109/L),with statistically significant difference(P＜0.05).There was no statistically significant difference in white blood cell(WBC)count and neutrophil(NEUT)percentage between two groups(both P＞0.05).Among the isolated Hin strains,27,2,and 2 strains were type b(Hib),e and f,respectively;3 strains were not typed;serotype a,c and d strains were not found.There was no statistically significant difference in the distribution of typeable Hin strains between the two groups(x2=0.25,P＞0.05).There was no statistically significant difference in the constituent rate of typeable Hin strains between male and fe-male children(67.74％vs 32.26％,x2=1.42,P＞0.05).Conclusion The majority of invasive Hin infection cases are children under 3 years old,and the predominant strain is type b.CRP and PCT levels of infected children in-creased significantly,while PLT is significantly lower than that of non-infected children,which has certain clinical diagnostic value and can provide effective support for early classified diagnosis and anti-infection treatment of inva-sive infectious diseases combined with other clinical testing items.

Objective To understand the distribution of ecological suitability of Scutellaria baicalensis Georgi;To screen the main ecological factors affecting its distribution;To predict its suitable planting area in China.Methods A total of 231 batches of Scutellaria baicalensis Georgi were collected through the fourth national survey of TCM resources.The environment information of sampling points were recorded,and 55 ecological factors were analyzed by MaxEnt model and GIS.Results Rainfall,vegetation type and soil types had a greater influence on the distribution of Scutellaria baicalensis Georgi.The ecological suitable areas of Scutellaria baicalensis Georgi mainly concentrated in the eastern part of Shaanxi Province,the western part of Hebei Province and most areas of Chengde,the most part of Shanxi Province,the western part of Beijing,the western part of Liaoning Province,the central and eastern part of Shandong Province,the junction of Jiangsu and Anhui Province,the northern part of Yunnan Province,and the eastern and southern part of Sichuan Province.Conclusion The ecological suitability areas of Scutellaria baicalensis Georgi were classified in this study,and the results could provide reference for the selection of cultivation areas of Scutellaria baicalensis Georgi.

Objective To establish a method for the simultaneous determination of aconitine,neoaconitine,hypaconitine,benzoyl aconitine,benzoyl mesaconine and benzoylhypacoitine in Xiaozhong ointment by UPLC-TQD-MS.Methods ACQUITY UPLC BEH C18 column(50 mm ×2.1 mm,1.7 μm),mobile phase 0.1％formic acid water(A)-acetonitrile(B),gradient elution,column temperature 40 ℃,flow rate 0.3 mL·min-1,injection volume 5 μL;electrospray ionization source(ESI+)and multiple reaction monitoring(MRM)were used for mass spectrometry analysis.Results The concentration of aconitine,new aconitine,hypaconitine,benzoyl aconitine,benzoyl new aconitine and benzoyl hypaconitine were 1.0-100.0 ng·mL-1,respectively,the average recovery were 98.62％-101.24％.The mass fractions of the six components were 0.18,0.33,0.38,0.43,0.28,0.06μg·g-1.Conclusion The method can be used to determine the content of 6 aconitine-type alkaloids in Xiaozhong ointment,and provide reference for the quality evaluation and clinical safe use of Xiaozhong ointment.

Cancer stem cells (CSCs), a small subset of cells in the tumor bulk with the ability of self-renewal and differentiation, are the key to tumor occurrence, metastasis, drug resistance and relapse. CSCs are resided in a specific microenvironment, and their number maintenance, self-renewal and differentiation are precisely regulated by the microenvironment, and the immune microenvironment is one of the most critical microenvironments for CSCs. In recent years, tumor immunotherapy has achieved great success, but drug resistance and recurrence are frequently occurred after immunotherapy. Compared with non-CSC tumor cells, CSCs harbor stronger immune escape ability, and their roles in tumor immune escape are increasingly followed. In this review, we described the discovery history and lineage sources of CSCs, focused on immune cells in the CSC microenvironment, such as tumor-infiltrating lymphocytes, tumor-associated macrophages, and tumor-associated dendritic cells, and analyzed the mechanism of CSC-immune cell interaction. Intervention strategies targeting CSCs and their immune microenvironment are also described. With the development and application of advanced technologies such as CSC-immune cell co-culture, single-cell sequencing and lineage tracing, the immune escape of CSCs can be suppressed by targeting the interaction between CSCs and immune cells or reversing the immunosuppressive microenvironment, which is expected to provide potential solutions to the problems of drug resistance and relapse in tumor immunotherapy.

Natural products are important sources of drug discovery. However, the traditional methods of extraction and isolation, as well as chemical synthesis for obtaining natural products are associated with issues such as operational complexity, high costs, low efficiency, and environmental pollution. Constructing microbial cell factories through synthetic biology methods to produce medicinal natural products has the advantages of high efficiency, low cost, and environmental protection. Nevertheless, the scope and yield improvement of the products are limited by the limitations of enzymes in microbial cell factories. Protein engineering is considered one of the most effective approaches to overcome these limitations. This article introduces commonly used methods of protein engineering technology and summarizes its specific applications in improving enzyme performance, modifying the enzymatic environment, and promoting the development of synthetic biology tools in the field of pharmaceutical natural product synthesis. Furthermore, it analyzes the current bottlenecks and challenges in protein engineering and looks forward to its future application prospects, offering insights for the development and practical use of protein engineering technology.

Intraperitoneal administration of timosaponin A-III (TA-III) has therapeutic effects on high-fat diet-induced metabolic dysfunction-associated steatotic liver disease (MASLD), but oral administration has no effect. This suggests that gut microbiota may affect the oral bioavailability of TA-III. Metabolic dysfunction-associated steatohepatitis (MASH) is an inflammatory subtype of MASLD. To investigate the therapeutic effect of different administration modes of TA-III on MASH and its relationship with gut microbiota metabolism. In this study, a MASH mouse model was induced by choline-deficient, L-amino acid-defined, high-fat diet (CDAHFD). Comparing the therapeutic effect of intraperitoneal injection (10 mg·kg^-1, ip) and intragastric administration (100 mg·kg^-1, ig) of TA-III. The concentration of TA-III in serum of rats under the two administration modes was analyzed. On this basis, the metabolic effect of gut microbiota on TA-III in mice was verified by the experiment of metabolism of gut microbiota in vitro. The pharmacokinetic experiment of combined antibiotic intervention in mice further verified the metabolism of TA-III by gut microbiota in mice. Finally, the concentration of TA-III in serum of mice after the administration of TA-III by intragastric administration under different antibiotic intervention conditions was compared, and 16S rRNA sequencing analysis was combined to find the key bacteria that may participate in the metabolism of TA-III. The animal welfare and experimental procedures in this paper were in accordance with the provisions of the Animal Ethics Committee of Shanghai University of Traditional Chinese Medicine. The ethics approval number is PZSHUTCM2307030004 and PZSHUTCM2310200003. The results showed that TA-III (10 mg·kg^-1, ip) had definite therapeutic effect on MASH mice, but TA-III (100 mg·kg^-1, ig) was ineffective. The analysis showed that the prototype concentration of TA-III in serum and liver of mice in TA-III (100 mg·kg^-1, ig) was significantly lower than TA-III (10 mg·kg^-1, ip), suggesting that the oral administration of TA-III may be metabolized by gut microbiota. The concentration of TA-III in serum of streptomycin (Str) treated mice was higher than normal mice. Combined with 16S rRNA gene sequencing analysis, it was found that the abundance of Akkermansia_muciniphila (A. muciniphila) was significantly reduced in the Str group. In vitro experiments showed that A. muciniphila could metabolize TA-III. In conclusion, gut microbiota is an important factor affecting the efficacy of TA-III administration through the gastrointestinal tract, in which A. muciniphila may play an important role.