Objective·To investigate the regulatory effects and underlying mechanisms of mouse mandibular osteoblasts on B cell differentiation and development.Methods·Single-cell suspensions from mouse mandibular bone were prepared using an optimized enzymatic digestion method and induced to differentiate into osteoblasts in vitro.Osteogenic potential was validated by real-time quantitative PCR(RT-qPCR),alkaline phosphatase(ALP)staining,and alizarin red S(ARS)staining.The spatial localization relationship between osteoblasts and B cells in mandibular tissues was examined via immunofluorescence staining.High-purity hematopoietic progenitor cells were isolated using fluorescence-activated cell sorting.A Transwell co-culture system was established to assess the regulatory effects of different osteoblast concentrations(5×104,2.5×105,and 5×105 cells/well)on B cell differentiation(5×104 cells/well).Flow cytometry and RT-qPCR were employed to evaluate B cell viability and differentiation.Additionally,RT-qPCR was used to analyze the expression of osteoblast-secreted factors associated with B cell development during osteogenic differentiation.Results·Mandibular osteoblasts exhibited robust osteogenic potential,as confirmed by ALP/ARS staining and high expression of osteogenic markers(Runx2,Osx,Ocn,and Alp)via RT-qPCR.Immunofluorescence revealed close spatial proximity between osteoblasts and B cells in mandibular tissues.In the co-culture system,osteoblasts promoted B cell differentiation in a concentration-dependent manner.RT-qPCR and immunofluorescence demonstrated that osteoblasts significantly upregulated key genes involved in B cell development(Ebf1,Rag1,Il7r,and Pax5;all P<0.001).Furthermore,osteoblast-derived factors(Il7,Baff,and Flt3l)were markedly elevated during osteogenic differentiation(all P<0.05).Conclusion·Mandibular osteoblasts enhance B cell differentiation and development in a concentration-dependent manner,likely through secreting growth factors that upregulate critical B cell differentiation genes.

Objective·To investigate the regulatory effects and underlying mechanisms of mouse mandibular osteoblasts on B cell differentiation and development.Methods·Single-cell suspensions from mouse mandibular bone were prepared using an optimized enzymatic digestion method and induced to differentiate into osteoblasts in vitro.Osteogenic potential was validated by real-time quantitative PCR(RT-qPCR),alkaline phosphatase(ALP)staining,and alizarin red S(ARS)staining.The spatial localization relationship between osteoblasts and B cells in mandibular tissues was examined via immunofluorescence staining.High-purity hematopoietic progenitor cells were isolated using fluorescence-activated cell sorting.A Transwell co-culture system was established to assess the regulatory effects of different osteoblast concentrations(5×104,2.5×105,and 5×105 cells/well)on B cell differentiation(5×104 cells/well).Flow cytometry and RT-qPCR were employed to evaluate B cell viability and differentiation.Additionally,RT-qPCR was used to analyze the expression of osteoblast-secreted factors associated with B cell development during osteogenic differentiation.Results·Mandibular osteoblasts exhibited robust osteogenic potential,as confirmed by ALP/ARS staining and high expression of osteogenic markers(Runx2,Osx,Ocn,and Alp)via RT-qPCR.Immunofluorescence revealed close spatial proximity between osteoblasts and B cells in mandibular tissues.In the co-culture system,osteoblasts promoted B cell differentiation in a concentration-dependent manner.RT-qPCR and immunofluorescence demonstrated that osteoblasts significantly upregulated key genes involved in B cell development(Ebf1,Rag1,Il7r,and Pax5;all P<0.001).Furthermore,osteoblast-derived factors(Il7,Baff,and Flt3l)were markedly elevated during osteogenic differentiation(all P<0.05).Conclusion·Mandibular osteoblasts enhance B cell differentiation and development in a concentration-dependent manner,likely through secreting growth factors that upregulate critical B cell differentiation genes.

Aim To investigate the inhibition role and mechanism of adipose derived mesenchymal stem cell(ADMSC)exosomes(Exo)on adverse ventricular remodeling after myocardial infarction(MI).Methods The chan-ges of autophagy and inflammasomes phenotype of cardiac fibroblasts after H2O2 treatment were observed.MI rats were in-jected with an equal volume of normal saline,adipose derived mesenchymal stem cell exosomes(MSC-Exo)or fibroblast exosomes(MEF-Exo)via a tail vein.The expression of autophagy related 16 like protein 1(ATG16L1),autophagy re-lated protein 7(ATG7)and NOD-like receptor protein 3(NLRP3),inflammatory response,the degree of myocardial fi-brosis,and the cardiac function were observed in different groups.Results After treatment with H2O2 on cardiac fi-broblasts,the expressions of ATG16L1 and ATG7 were significantly decreased(P＜0.001),NLRP3 was significantly in-creased(P＜0.001),and the levels of inflammatory cytokines interleukin-1β(IL-1β)and IL-18 were significantly elevated(P＜0.001).After MI rats were intervened with MSC-Exo,the expressions of autophagy related proteins ATG16L1 and ATG7 were significantly up-regulated(P＜0.001),NLRP3 was significantly down-regulated(P＜0.001),serum IL-1β and IL-18 levels were significantly decreased(P＜0.001),fibrosis-related proteins collagen Ⅰ and Ⅲ were significantly reduced(P＜0.001),myocardial fibrosis was significantly relieved(P＜0.001),and cardiac function was sig-nificantly improved(P＜0.001).Conclusion Adipose derived MSC-Exo play a role in inhibiting adverse ventricular remodeling after MI by regulating the balance of autophagy and NLRP3 inflammasomes.

Chronic refractory wounds have been a challenge for clinical treatment because of their diverse causative factors and complex pathological processes, long healing times, and high treatment costs. The microenvironment of the wound surface includes the external microenvironment of the periwound surface, the internal microenvironment of the wound surface, and subsurface physiological structures. Research on clinical treatment strategies based on the microenvironment of chronic refractory wounds continues to innovate and make progress. Hydrogels have the advantages of high-water content, adjustable performance, good biocompatibility, and similarity to extracellular matrix. The ability of hydrogels to load drugs and their modification to confer excellent tissue adhesion, antibacterial, antioxidant, and modulation of inflammatory factor expression can be used to achieve a multi-factor response and modulation of the physical, chemical, and biological aspects of the trauma microenvironment. Therefore, hydrogels have outstanding advantages and clinical application prospects in the repair of chronic, difficult-to-heal wounds. In this review paper, the characteristics and etiology of chronic refractory wounds were introduced, and the classification of microenvironment-responsive hydrogels for chronic refractory wounds and their application in the repair of refractory wounds were reviewed. Besides, the shortcomings of current hydrogels were discussed, and an outlook was proposed.

In the original publication, the label of Fig. 2C should be read as "GFAP/lectin/DAPI" not "DMP1/GFAP/lectin/DAPI".

The blood-brain barrier (BBB) is a tight boundary formed between endothelial cells and astrocytes, which separates and protects brain from most pathogens as well as neural toxins in circulation. However, detailed molecular players involved in formation of BBB are not completely known. Dentin matrix protein 1 (DMP1)-proteoglycan (PG), which is known to be involved in mineralization of bones and dentin, is also expressed in soft tissues including brain with unknown functions. In the present study, we reported that DMP1-PG was expressed in brain astrocytes and enriched in BBB units. The only glycosylation site of DMP1 is serine89 (S89) in the N-terminal domain of the protein in mouse. Mutant mice with DMP1 point mutations changing S89 to glycine (S89G), which completely eradicated glycosylation of the protein, demonstrated severe BBB disruption. Another breed of DMP1 mutant mice, which lacked the C-terminal domain of DMP1, manifested normal BBB function. The polarity of S89G-DMP1 astrocytes was disrupted and cell-cell adhesion was loosened. Through a battery of analyses, we found that DMP1 glycosylation was critically required for astrocyte maturation both in vitro and in vivo. S89G-DMP1 mutant astrocytes failed to express aquaporin 4 and had reduced laminin and ZO1 expression, which resulted in disruption of BBB. Interestingly, overexpression of wild-type DMP1-PG in mouse brain driven by the nestin promoter elevated laminin and ZO1 expression beyond wild type levels and could effectively resisted intravenous mannitol-induced BBB reversible opening. Taken together, our study not only revealed a novel element, i.e., DMP1-PG, that regulated BBB formation, but also assigned a new function to DMP1-PG.
Animals ; Astrocytes ; cytology ; metabolism ; Blood-Brain Barrier ; cytology ; metabolism ; Cells, Cultured ; Extracellular Matrix Proteins ; genetics ; metabolism ; Female ; Glycosylation ; Male ; Mice ; Proteoglycans ; metabolism ; Reverse Transcriptase Polymerase Chain Reaction

Objective To establish an auditory evoked potential (AEP) detection system in zerbafish.Methods The AEP detection tank was designed and made, and then verified for its quality and reliability via four experiments: anesthesia experiment, swim bladder deflation, noise exposure and goldfish AEP test.Finally, zebrafish (total length form 10 mm to 46 mm) were determined using this system for AEP.Zebrafish randomly were divided into five groups according to total length (TL=12~15 mm, n=6;TL=17~20 mm,n=4;TL=22~26 mm, n=4;TL=32~37 mm, n=9;and TL=42~46 mm,n=12).Goldfish, as control group, were purchased for local petshop (TL=38~54 mm,n=8).Results The results of these four verifying experiments confirmed the biological, rather than artefactual, nature of the responses represented by the recorded waveforms.The AEPs were detected up to a much higher frequency limit (12 kHz) than previously reported.In this study, all fish demonstrated a range of hearing frequency from 100 Hz to 12 kHz without frequency expansion during development.The best hearing was observed at 600 Hz~1 kHz.The mean values of the frequency-averaged thresholds (mean SEM) were 141.7±1.32, 124.8±1.31, 121.8±1.49, 117.8 ±1.09 and 124.4±1.87 dB w, respectively, for the 5 TL groups.The AEP thresholds demonstrated both developmental improvement and age-related loss of hearing sensitivity.Conclusion An auditory evoked potential detection system of zerbafish has been established with stable performance and can be used for AEP detection of zebrafish.

Objective To investigate the effects of different doses of diphtheria toxin on cochlear structure and auditory function of adult wildtype mice.Methods The auditory-mature wild type C57BL/6J mice 4 weeks old were randomly devided into 50 ng/g group, 100 ng/g group and control group.C57BL/6J mice in the 50 ng/g or 100 ng/g group were injected 50 ng/g or 100 ng/g diphtheria toxin intraperitoneally for one time, respectively, and the control mice were injected equal volume of normal saline for one time.Then we investigated the ABR threshold change and morphological change of inner and outer hair cell and spiral ganglion neuron 7 days after the injection.Results At 7 day post diphtheria toxin injection compared with those of in control group, in the 50 ng/g group, there was no threshold elevation across frequencies(8 kHz ABR threshold was 20.0±3.78 and 20.83±2.04 dB SPL for 50 ng/g and control respectively), and no loss of inner and outer hair cells (for both groups, the HC loss rates were 0.3%~1%) or SGN (the SGN density was 39.45±3.65, 41.03±3.73/105 μm2, in 50 ng/g and control, respectively).However, the 100 ng/g group, compared with those of in control group, the ABR threshold (8 kHz ABR threshold was 63.0±4.47 dB SPL, respectively)was significantly elevated across each frequency(t=19.62,P<0.001), and there was significant loss of outer hair cell (the loss rate of IHC and OHC was 0.5%±0.1%, 10.7%±0.3%, respectively), which was 10% loss in the apical, middle and basal turn(t=42.219,P<0.001).And the loss of spiral ganglion neuron (the SGN density was 25.55±3.66/105 μm2) was 38%, which was significantly different from the control(t=10.985,P<0.001).Conclusion High dose injection of diphtheria toxin can cause loss of outer hair cell and spiral ganglion neuron in wild type auditory-mature C57BL/6J mice.

Binding Sites ; Histone-Lysine N-Methyltransferase ; chemistry ; metabolism ; Histones ; chemistry ; metabolism ; Humans ; Methylation ; Multiprotein Complexes ; chemistry ; metabolism ; Nuclear Proteins ; chemistry ; metabolism ; Protein Binding ; Protein Conformation ; Yeasts ; chemistry

Objective To compare the efficacy of three solutions used in the patients who suffered chronic obstructive pulmonary diseases (COPD ) when combined with oral fungal infection,and to find the most suitable one. Methods Using randomized controlled trial method, 98 patients who suffered COPD complicated with fungal infection were randomly divided into three groups. Method 1, using 0.5% povidone for the oral care. Method 2, making 200 million U of nystatin powder dissolved in 2.5% NaHCO3 for the oral care.Method 3, gave oral care with 2.5% NaHCO3 first, then put 200 million U of nystatin powder into 20 ml glycerol and covered the oral spots with them. Three groups were observed in the effect of oral fungal infection and the feedback from patients and nurses. Results In the analysis of the white spots sharply disappeared or pharyngeal swab smear turned negative, method 1 and method 3 had no statistical difference, P>0.05, but method 2 and method 3 had statistical significance, t=3.892, P <0.01; the three groups had no significant difference of feedback from the patients’ oral comfort, P >0.05. The time made solution of method 1 occupied 41.7% of method 3, and the time made solution of method 2 occupied 54.4%of method 3, while there was no statistical significance in total time of three groups (P>0.05). Conclusions The oral care with 0.5% povidone had good effect on the patients who suffered COPD complicated with oral fungal infection,with good comfort,and most nurses saving time. Nystatin powder dissolved in 2.5% NaHCO3 worked later than nystatin powder dissolved in glycerol.