Nanoparticles (NPs)-mediated Siglec15 silencing and macrophage repolarization for enhanced cancer immunotherapy

T cell infiltration and proliferation in tumor tissues are the main factors that significantly affect the therapeutic outcomes of cancer immunotherapy. Emerging evidence has shown that interferon-gamma (IFNγ) could enhance CXCL9 secretion from macrophages to recruit T cells, but Siglec15 expressed on TAMs can attenuate T cell proliferation. Therefore, targeted regulation of macrophage function could be a promising strategy to enhance cancer immunotherapy via concurrently promoting the infiltration and proliferation of T cells in tumor tissues. We herein developed reduction-responsive nanoparticles (NPs) made with poly (disulfide amide) (PDSA) and lipid-poly (ethylene glycol) (lipid-PEG) for systemic delivery of Siglec15 siRNA (siSiglec15) and IFNγ for enhanced cancer immunotherapy. After intravenous administration, these cargo-loaded could highly accumulate in the tumor tissues and be efficiently internalized by tumor-associated macrophages (TAMs). With the highly concentrated glutathione (GSH) in the cytoplasm to destroy the nanostructure, the loaded IFNγ and siSiglec15 could be rapidly released, which could respectively repolarize macrophage phenotype to enhance CXCL9 secretion for T cell infiltration and silence Siglec15 expression to promote T cell proliferation, leading to significant inhibition of hepatocellular carcinoma (HCC) growth when combining with the immune checkpoint inhibitor. The strategy developed herein could be used as an effective tool to enhance cancer immunotherapy.

Clinical feature of 39 patients with acute brucellosis from Shenzhen: results of a retrospective analysis / 中华地方病学杂志

Objective To analyze the clinical features of patients with acute stage brucellosis in Shenzhen,and provide a scientific basis for prevention and control of brucellosis in immigrant city.Methods A retrospective analysis was conducted to collect clinical data of patients with brucellosis admitted to the Department of Infectious Diseases,Shenzhen People's Hospital from May 2013 to May 2018.The patient's epidemiology manifestations,pathogen and laboratory examination results,diagnosis and treatment outcomes and prognosis were analyzed.Results Among the 39 patients with brucellosis,males were predominant,with a male to female ratio of 1.4 ∶ 1.0 (23 ∶ 16),an age of (44.91 ± 17.18) years and 24 cases were non-Guangdong natives.There were 23 cases with epidemiological history,including 14 cases with mutton,sheep viscera and goat milk history;the disease occurred throughout the year,mainly from February to July,a total of 26 cases.The clinical manifestations of the patients were mainly fever,sweating,fatigue,joint and muscle pain,weight loss,and liver or spleen or lymph nodes swelling.The blood culture was identified as 38 cases of Brucella melitensis and 1 case of Brucella suis.All strains were sensitive to common antibiotics in vitro.All cases were diagnosed as acute stage of brucellosis,2 cases with orchitis,1 case with brucellosis meningoencephalitis,3 cases with spondylitis,and 3 cases with misdiagnosis.Thirty-nine patients were cured according to the "Brucellosis Diagnosis and Treatment Guidelines (Trial)" and were followed up for 1 year.Conclusions Patients with brucellosis in Shenzhen are mainly infected with Brucella melitensis;fever,sweating,joint and muscle pain are the main clinical symptoms;the patient's efficacy and prognosis are better after treatment;for the occurrence of occasional misdiagnosis,it is recommended that in immigrant cities,medical staff should strengthen their understanding of brucellosis.

An analysis of epidemiological and clinical features of 46 patients with hemorrhagic fever with renal syndrome in Shenzhen City / 中华地方病学杂志

Objective@#To understand the epidemiological and clinical features of patients with hemorrhagic fever and renal syndrome (HFRS) in Shenzhen, and to accumulate experience in the diagnosis and treatment of HFRS in this area.@*Methods@#A retrospective analysis was conducted by collecting the clinical data from 46 patients who were confirmed with HFRS and admitted to the Department of Infectious Diseases of Shenzhen People's Hospital from January 2015 to December 2018. The demographic characteristics, epidemiological, clinical manifestations, examinations, treatments and prognosis, and other characteristics were analyzed.@*Results@#All the 46 patients with HFRS were residens in Shenzhen, with a male-to-female ratio of 6.67∶1.00(40∶6), aged (40.18 ± 15.63) years old, and 38 patients (82.61%) aged 23-45 years old. There were 41 patients (89.13%) with a history of HFRS epidemiology, and there were mice in their houses or workplaces. The houses of 39 patients (84.78%) were rented, and 34 patients(87.18%) rented their houses in urban villages. There were morbidity throughout the year, and 33 patients (71.74%) were ill from January to June. In clinical classification, 44 cases (95.65%) were mild, 2 cases (4.35%) were medium, and there were no severe or critical cases. The clinical manifestations were that all patients were hospitalized due to fever mainly with hyperthermia. Thirty-nine patients (84.78%) were presented with systemic aches, headaches, low back pain and eyelid pain, and 28 patients (60.87%) had skin and mucous membrane hyperemia flushing. Clinical stages showed that all patients had pyretogenesis stage and polyuria stage, including pyretogenesis stage [(7.34 ± 6.82) d], polyuria stage [(9.94 ± 5.77) d], only 4.35% (2/46) patients with hypotension shock stage, all patients did not have oliguric stage. On the next day of admission, the number of white blood cells in 46 patients was (8.17 ± 3.19) × 109/L, and 38 cases (82.61%) in the normal range; platelet was (61.92 ± 32.53) × 109/L, and 42 cases (91.30%) were decreased; the procalcitonin was (1.62 ± 0.38) ng/ml, and 41 cases (89.13%) were increased; C-reactive protein was (74.33 ± 30.48) mg/L, and 46 patients (100.00%) were elevated; creatinine was (176.25 ± 55.15) μmol/L, and 19 cases (41.30%) were increased. Abnormal liver function was manifested by increased enzymology, alanine aminotransferase was (137.58 ± 46.76) U/L, and aspartate aminotransferase was (129.82 ± 40.29) U/L. All patients were positive for Hantavirus IgM. B-ultrasound results showed that in 58.70% (27/46) patients, both kidneys were plump and the parenchymal echo was enhanced. Liver injury occurred in 39 patients (84.78%) and sinus bradycardia in 2 patients (4.35%). All the 46 patients were clinically cured.@*Conclusion@#The epidemiological characteristics of patients with HFRS in Shenzhen are typical, the clinical manifestations are mild, and the curative effect and prognosis are good.

An analysis of epidemiological and clinical features of 46 patients with hemorrhagic fever with renal syndrome in Shenzhen City / 中华地方病学杂志

Objective To understand the epidemiological and clinical features of patients with hemorrhagic fever and renal syndrome (HFRS) in Shenzhen,and to accumulate experience in the diagnosis and treatment of HFRS in this area.Methods A retrospective analysis was conducted by collecting the clinical data from 46 patients who were confirmed with HFRS and admitted to the Department of Infectious Diseases of Shenzhen People's Hospital from January 2015 to December 2018.The demographic characteristics,epidemiological,clinical manifestations,examinations,treatments and prognosis,and other characteristics were analyzed.Results All the 46 patients with HFRS were residens in Shenzhen,with a male-to-female ratio of 6.67:1.00 (40:6),aged (40.18 ± 15.63) years old,and 38 patients (82.61％) aged 23-45 years old.There were 41 patients (89.13％) with a history of HFRS epidemiology,and there were mice in their houses or workplaces.The houses of 39 patients (84.78％) were rented,and 34 patients (87.18％) rented their houses in urban villages.There were morbidity throughout the year,and 33 patients (71.74％) were ill from January to June.In clinical classification,44 cases (95.65％) were mild,2 cases (4.35％) were medium,and there were no severe or critical cases.The clinical manifestations were that all patients were hospitalized due to fever mainly with hyperthermia.Thirty-nine patients (84.78％) were presented with systemic aches,headaches,low back pain and eyelid pain,and 28 patients (60.87％) had skin and mucous membrane hyperemia flushing.Clinical stages showed that all patients had pyretogenesis stage and polyuria stage,including pyretogenesis stage [(7.34 ± 6.82) d],polyuria stage [(9.94 ± 5.77) d],only 4.35％ (2/46) patients with hypotension shock stage,all patients did not have oliguric stage.On the next day of admission,the number of white blood cells in 46 patients was (8.17 ± 3.19) × 109/L,and 38 cases (82.61％) in the normal range;platelet was (61.92 ±32.53) × 109/L,and 42 cases (91.30％) were decreased;the procalcitonin was (1.62 ± 0.38) ng/ml,and 41 cases (89.13％) were increased;C-reactive protein was (74.33 ± 30.48) mg/L,and 46 patients (100.00％) were elevated;creatinine was (176.25 ± 55.15) μmol/L,and 19 cases (41.30％) were increased.Abnormal liver function was manifested by increased enzymology,alanine aminotransferase was (137.58 ± 46.76) U/L,and aspartate aminotransferase was (129.82 ± 40.29) U/L.All patients were positive for Hantavirus IgM.B-ultrasound results showed that in 58.70％ (27/46) patients,both kidneys were plump and the parenchymal echo was enhanced.Liver injury occurred in 39 patients (84.78％) and sinus bradycardia in 2 patients (4.35％).All the 46 patients were clinically cured.Conclusion The epidemiological characteristics of patients with HFRS in Shenzhen are typical,the clinical manifestations are mild,and the curative effect and prognosis are good.

Bioinformatic analysis of streptococcus mutans thrS gene and construction of homologous recombinant plasmids / 中国组织工程研究

BACKGROUND: Streptococcus mutans (S. mutans) is generally considered to be the principal aetiological agent for dental caries, thrS gene may relate to the virulence of S. mutans involved in the adherence, acidogenicity and acidodurance.OBJECTIVE: To investigate the conservation status of the thrS gene of S. mutans and to construct the homologous recombinant plasmid.METHODS: Southern Blot was used to analyze the distdbuUon of thrS gene in S. mutans. The upstream and downstream sequences of thrS gene were cloned respectively into multiple cloning sites of suicide plasmid pFW5 to construct the recombinant plasmid,RESULTS AND CONCLUSION: The thrS gene was conserved in 6 strains of S. mutans in this test. By PCR analysis and enzyme digesting, it was confirmed that S. mutans thrS gene homologous recombinant plasmid was successfully constructed,which can be used in future research of construction of thrS -negative mutans of S. mutans strain UA159.

Construction of psm-negative mutans of S.mutans UA159 / 实用口腔医学杂志

Objective:To knock the phospho-sugar mutase gene (psm) out of the genome of S. mutans strain UA159 and to construct the mutant will lay a foundation for the study of the function of the psm. Methods:Two upstream and downstream DNA sequences of the psm were selected and cloned respectively into multiple cloning sites I and II of suicide plasmid pFW5 to construct the recombinant plasmid which was confirmed by enzyme digesting and sequencing. The recombinant plasmid was transformed into S. mutans UA159 by natural transformation and antibiotic was used to screen the positive transformants. According to the principle of homologous recombination, allelic exchange between the recombinant plasmid and S. mutans UA159 was achieved. Results:By PCR analysis and sequencing, it was confirmed that the psm of S. mutans UA159 was substituted for resistant gene of spectinomycin. Conclusion: The psm-negative mutants of S. mutans UA159 is successfully constructed.

High-level expression and purification of ComE in Escherichia coli / 实用口腔医学杂志

Objective:To construct the prokaryotic expression vector pET28a(+)-comE, to get the highest expression of comE in E. coli BL21(DE3), and to obtain the purified ComE fusion protein. Methods: ComE gene was amplified by PCR with specific primers from genome of Streptococcus mutans UA159. After digested by two restriction endonucleases BamH Ⅰand Xho Ⅰ, the gene segment was inserted into vector pET28a(+). Then the recombinant vector was transformed into E. coli BL21(DE3). The protein expression was induced by IPTG and the result was confirmed by Western blot. The solubility of the fusion protein was examined by 12% SDS-PAGE, and the expression conditions were optimized. Finally the fusion protein was purified by a two-step purification procedure utilizing Ni2+ chelating column and S75 size exclusion column. Results: The recombinant plasmid was confirmed by PCR, restriction endonucleases digestion and sequence analysis, which showed that the inserted segment was identical to comE gene reported by GenBank without nucleotide mutation. The fusion protein which was confirmed by Western blot can be expressed in E. coli BL21(DE3) and existed both in supernatant and precipitation. The maximum expression product amount was obtained when the A600 value of E. coli BL21(DE3) was 0.4, IPTG concentration was 0.10 mmol/L and induction time was 4 h. Conclusion: The recombinant plasmid is constructed and the ComE fusion protein is purified by the optimum conditions.