A hybrid bacterium with tumor-associated macrophage polarization for enhanced photothermal-immunotherapy

Remodeling the tumor microenvironment through reprogramming tumor-associated macrophages (TAMs) and increasing the immunogenicity of tumors via immunogenic cell death (ICD) have been emerging as promising anticancer immunotherapy strategies. However, the heterogeneous distribution of TAMs in tumor tissues and the heterogeneity of the tumor cells make the immune activation challenging. To overcome these dilemmas, a hybrid bacterium with tumor targeting and penetration, TAM polarization, and photothermal conversion capabilities is developed for improving antitumor immunotherapy in vivo. The hybrid bacteria (B.b@QDs) are prepared by loading Ag2S quantum dots (QDs) on the Bifidobacterium bifidum (B.b) through electrostatic interactions. The hybrid bacteria with hypoxia targeting ability can effectively accumulate and penetrate the tumor tissues, enabling the B.b to fully contact with the TAMs and mediate their polarization toward M1 phenotype to reverse the immunosuppressive tumor microenvironment. It also enables to overcome the intratumoral heterogeneity and obtain abundant tumor-associated antigens by coupling tumor penetration of the B.b with photothermal effect of the QDs, resulting in an enhanced immune effect. This strategy that combines B.b-triggered TAM polarization and QD-induced ICD achieved a remarkable inhibition of tumor growth in orthotopic breast cancer.

Avaliação do efeito de cepas probióticas em biofilme de S. aureus sobre discos de titânio com superfície tratada / Assessment of probiotic bacterial strains effect on S. aureus biofilm on titanium discs with treated surfaces

Resumo Introdução As doenças peri-implantares apresentam como um dos principais fatores etiológicos o biofilme bacteriano, geralmente formado por uma microbiota semelhante à das doenças periodontais. Seu tratamento está centrado na descontaminação da superfície do implante e na remoção mecânica do biofilme, podendo ainda estar associado à administração de agentes antimicrobianos. Nesse sentido, tem sido cogitada a utilização de probióticos, que são microrganismos benéficos à saúde e que podem ter grande importância na cavidade oral, como coadjuvante no tratamento das peri-implantites. Objetivo Avaliar o efeito das cepas probióticas de Lactobacillus brevis e Bifidobacterium bifidum no crescimento do biofilme monoespécie de Staphylococcus aureus. Material e método Discos de titânio padronizados e com superfície tratada foram submersos em meio contendo caldo BHI e Staphylococcus aureus durante sete dias. Após esse período, o caldo foi retirado, os discos foram lavados e, então, introduzidos em um novo caldo BHI contendo as suspensões probióticas, sendo assim comparados a um grupo controle, sem probióticos. As amostras foram incubadas por 24h e então foram realizadas as diluições e a contagem das UFC (unidades formadoras de colônia) para Staphylococcus aureus. Resultado Após análise estatística dos dados, observou-se que a adição de ambos os probióticos resultaram em redução significativa (p<0,05) de UFC, quando comparados ao controle. Conclusão Conclui-se que os probióticos analisados (Lactobacillus brevis e Bifidobacterium bifidum) reduziram consideravelmente o crescimento do patógeno Staphylococcus aureus. Além disso, a cepa de Lactobacillus brevis apresentou efeito inibidor superior ao da cepa Bifidobacterium bifidum para ser utilizada como controle do biofilme bacteriano de Staphylococcus aureus.

Abstract Introduction One of the main etiological factors for peri-implant diseases is the bacterial biofilm, which usually features a similar microbiota to periodontal diseases. Its treatment focus on the decontamination of the implant surface and on the mechanical removal of biofilm, and it may also be associated to the administration of antimicrobial agents. Thus, the use of probiotics has been considered, since they feature beneficial microorganisms to health and may be of great importance for the oral cavity as an adjunct for the treatment of peri-implant diseases. Objective The aim of this in vitro study was to assess the effect of probiotic strains of Lactobacillus brevis and Bifidobacterium bifidum on the growth of single-species biofilm of Staphylococcus aureus. Material and method Standardized surface-treated titanium discs were submerged in a medium containing BHI broth and Staphylococcus aureus, for 7 days. After this period, the broth was removed, the discs were washed and, then, submerged in a new BHI broth containing probiotic suspensions and compared to a control group (with no probiotics). Samples were incubated for 24 hours and then the dilutions and CFU (colony-forming units) counting for Staphylococcus aureus were performed. Result Statistical analysis revealed that the addition of both probiotics resulted in a significant reduction (p<0,05) of CFU, when compared to the control group. Conclusion The assessed probiotics (Lactobacillus brevis and Bifidobacterium bifidum) considerably reduced Staphylococcus aureus growth. In addition, Lactobacillus brevis strain presented a superior inhibition effect than Bifidobacterium bifidum strain for Staphylococcus aureus bacterial biofilm control.

Construction and expression of a recombinant Bifidobacterium bifidum vaccine of Schistosoma japonicum / 中华地方病学杂志

Objective To construct a recombinant Bifidobacterium bifidum (Bb)vaccine[Bb (pGEX-Sj26GST-Sj14-3-3)] of Schistosomajaponicum (Sj) and analyze the expression of the fusion gene Sj26GST-Sj14-3-3 of Sj in Bb.Methods The recombinant plasmid pGEX-Sj26GST-Sj14-3-3 was electroporated into Bb to construct a recombinant Bb (pGEX-Sj26GST-Sj14-3-3) vaccine.Mter induction with isopropyl-β-D-thiogalactoside (IPTG),double restriction enzymes digestion and polymerase chain reaction (PCR) were used to identify the recombinant Bb (pGEX-Sj26GST-Sj14-3-3),expression of the recombinant protein was analyzed and identified by sodium dodecyl sulfonate-polyacrylamide gel electrophoresis (SDS-PAGE) and Western blotting.Results The recombinant plasmid pGEX-Sj26GST-Sj14-3-3 was successfully transformed into Bb identified by double restriction enzymes digestion and PCR.SDS-PAGE analysis showed that the relative molecular mass of the expressed recombinant protein was approximately 67 × 103.The expressed protein could be recognized by the immune sera from rabbits infected with Sj by Western blotting.Conclusions The recombinant Bb (pGEX-Sj26GST-Sj14-3-3) vaccine of Sj is successfully constructed.The fusion gene Sj26GST-Sj14-3-3 can be expressed in recombinant Bb and the expressed target protein shows specific antigenicity.

Construction and identification of recombinant vaccine Bifidobacterium bifidum (pGEX-Sj32) of Schistosoma japonicum / 中华地方病学杂志

Objective To construct and identify recombinant vaccine Bifidobacterium bifidum (pGEX-Sj32) of Schistosomajaponicum (Sj).Methods The Sj32 gene amplified by PCR from template of plasmid pET28α-Sj32 that extracted from recombinant Escherichia coli(E.coli) BL21 (pET28α-Sj32) stored in our laboratory was cloned into E.coli-Bifidobacterium bifidum shuttle expression vector pGEX-1λT to construct recombinant plasmid pGEX-Sj32.The recombinant plasmid was transformed into E.coli BL21 (DE3).The plasmid was extracted and identified by restriction enzyme digestion.The recombinant Bifibacterium bifidum (pGEX-Sj32) vaccine was constructed by electroporating pGEX-Sj32 into Bb.The extracted plasmid was amplified and identified by PCR.Results The gene Sj32 of 1 270 bp in length was amplified by PCR.The restriction endonuclease digestion showed that the length of plasmid vector was 4 947 bp,and Sj32 gene was 1 270 bp.The Sj32 gene amplified by PCR from the template of pGEX-Sj32 extracted from recombinant Bb (pGEX-Sj32) vaccine was 1 270 bp in length which consistent with expected result.Conclusion The recombinant Bb (pGEX-Sj32) vaccine of Sj is successfully constructed.

Growth Characteristics Modeling of Mixed Culture of Bifidobacterium bifidum and Lactobacillus acidophilus using Response Surface Methodology and Artificial Neural Network

Different culture conditions viz. additional carbon and nitrogen content, inoculum size and age, temperature and pH of the mixed culture of Bifidobacterium bifidum and Lactobacillus acidophilus were optimized using response surface methodology (RSM) and artificial neural network (ANN). Kinetic growth models were fitted for the cultivations using a Fractional Factorial (FF) design experiments for different variables. This novel concept of combining the optimization and modeling presented different optimal conditions for the mixture of B. bifidum and L. acidophilus growth from their one variable at-a-time (OVAT) optimization study. Through these statistical tools, the product yield (cell mass) of the mixture of B. bifidum and L. acidophilus was increased. Regression coefficients (R2) of both the statistical tools predicted that ANN was better than RSM and the regression equation was solved with the help of genetic algorithms (GA). The normalized percentage mean squared error obtained from the ANN and RSM models were 0.08 and 0.3%, respectively. The optimum conditions for the maximum biomass yield were at temperature 38°C, pH 6.5, inoculum volume 1.60 mL, inoculum age 30 h, carbon content 42.31% (w/v), and nitrogen content 14.20% (w/v). The results demonstrated a higher prediction accuracy of ANN compared to RSM.

Curative effect and safety of the adjuvant therapy of bifidobacterium bifidum-sine capsules to helicobacter pylori positive peptic ulcer / 中国基层医药

Objective To discuss the curative effect and safety of the adjuvant therapy ( AT) of bifidobacteri-um bifidum-sine capsules to helicobacter pylori ( HP) positive peptic ulcer .Methods 76 HP infected peptic ulcer patients were selected and divided into the observation group and control group at random .The patients in control group were given traditional triple therapy of omeprazole ,amoxicillin and carat enzyme for 4 weeks,while the patients in observation group were given bifidobacterium bifidum-sine capsules(420mg) three times daily for 4 weeks.Clinical symptom remission,ulcer healing and HP clearance of patients in two groups were observed and compared .The inci-dence rate of drug adverse reaction ( DAR) during treatment period was observed ,and the recurrence rates of peptic ulcer within half a year and one year were followed up .Results The HP clearance rate of the observation group was significantly higher than control group (χ2 =4.21,P<0.05).The incidence rate of DAR of the observation group was 10.26%,which was significantly lower than 29.73%of the control group (χ2 =4.55,P<0.05).34 cases in the ob-servation group and 30 cases in the control group were healed ,these patients were followed up for half a year and one year,and the recurrence rate of peptic ulcer in observation group was significantly lower than that in control group (χ2 =4.51,5.23,all P<0.05).Conclusion The bifidobacterium bifidum-sine capsules combined with PPI triple therapy,can increase the eradication rate of HP ,reduce DAR and the recurrence rate of ulcer .

Dynamic observation of splenocyte apoptosis in mice immunized with recombinant vaccine Bifidobacterium bifidum pGEX-Sj14-3-3 of Schistosoma japonicum / 中国地方病学杂志

Objective To investigate the effects of recombinant vaccine Bifidobacterium bifidum (Bb) pGEX-Sj14-3-3 on splenocyte apoptosis in BALB/c mice.Methods Ninety-six BALB/c mice were randomly divided into two groups according to their body mass:per os group (PO) and intranasal immunization group (IN),with 48 mice in each group.All mice were orally and intranasally immunized with recombinant vaccine Bb(pGEX-Sj14-3-3).Four mice in each group were sacrificed on weeks 0,2,4,6,8,10,12,14,16,18,20 and 22,respectively,after immunization,and splenocytes were separated and cultured with or without ConA stimulation.The apoptotic rates of splenocytes were detected by flow cytometry.Results It showed that apoptotic level of splenocytes in both groups remarkably increased after 2-4 weeks without ConA stimulation (PO:0.069 ± 0.005,0.076 ± 0.010; IN:0.037 ± 0.002,0.075 ± 0.002),and the value reached the peak on the 4th week,and the differences were statistically significant compared with that of week 0(all P ＜ 0.05).Apoptotic level of splenocytes in both groups with ConA stimulation increased after 2-6 weeks(PO:0.089± 0.006,0.098 ± 0.010,0.060±0.007; IN:0.054 ± 0.001,0.093 ± 0.003,0.058 ± 0.012),and the value also reached the peak after 4 week,respectively.The differences were statistically significant compared with that of week 0 (all P ＜ 0.05).Apoptotic level of splenocytes in each group with ConA stimulation was significantly higher than that without ConA stimulation.Conclusion It is suspected that the recombinant vaccine Bb(pGEX-Sj14-3-3) may inhibit apoptosis of splenocytes in mice immunized orally or intranasally.

Construction and identification of recombinant vaccine Bifidobacterium bifidum pGEX-Sj14-3-3 of Schistosoma japonicum / 中国地方病学杂志

Objective To construct and identify recombinant vaccine Bifwlobacterium bifidum(Bb)pGEX-Sj14-3-3 of Schistosoma japonicum(Sj). Methods Total RNA was extracted from adult Sj, antigen encoding gene Sj14-3-3 was amplified by RT-PCR and cloned into Escherichia coli (E. coli)-Bb shuttle expression vector pGEX-1λT to construct recombinant plasmid pGEX-Sj14-3-3. The recombinant plasmid was transformed into E. coli BL21 (DE3).The plasmid was extracted and identified by using BamH I and EcoR I. Then pGEX-Sjl4-3-3 was electroporated into Bb to construct recombinant Bb (pGEX-Sj14-3-3) vaccine. The extracted plasmid of the recombinant Bb (pGEX-Sj14-3-3) vaccine was identified by PCR, and the size of the products was compared with Sj14-3-3 gene of adult worms.Results Sj14-3-3 of 399 bp in length was amplified by RT-PCR. The products were digested by BamH I and EcoR I , and the fragments length of plasmid pGEX-Sj14-3-3 vector was 4947 bp, and of Sj 14-3-3 gene was 399 bp.The product of 399 bp Sj14-3-3 gene was also amplified by PCR from template of the extracted plasmid of the recombinant Bb(pGEX-Sj14-3-3 ) vaccine. The size of the product obtained was just the same as expected.Conclusion The recombinant Bb(pGEX-Sj14-3-3) vaccine of Sj is successfully constructed.

Effect of Probiotics on Symptoms in Korean Adults with Irritable Bowel Syndrome

BACKGROUND/AIMS: Irritable bowel syndrome (IBS) is a troublesome disease. Some strains of probiotics reportedly exert remarkable immunomodulatory effects, and so we designed a prospective double-blind randomized placebo-controlled clinical study to assess their effects in Korean adults with IBS. METHODS: IBS patients who met Rome III criteria were randomly assigned to receive composite probiotics or placebo. A total of 20 billion lyophilized bacteria were administered twice daily for 8 weeks. Primary outcome variables were symptom scores consisting of abdominal pain, flatulence, defecation discomfort, and sum of symptom scores. A visual analogue scale was used to quantify the severity. Secondary outcome variables consisted of the quality of life and bowel habits including defecation frequency and stool form. RESULTS: Thirty-six and 34 patients were randomized to the probiotics and placebo groups, respectively. Intention- to-treat analysis showed significant reductions in pain after 8 weeks of treatment: -31.9 and -17.7 in the probiotics and placebo groups, respectively (p=0.045). The reductions in abdominal pain, defecation discomfort, and sum of scores were more significant in 58 patients with a score of at least 3 on the baseline stool-form scale. CONCLUSIONS: Composite probiotics containing Bifidobacterium bifidum BGN4, Lactobacillus acidophilus AD031, and other species are safe and effective, especially in patients who excrete normal or loose stools.