Journal: Toxins

Article Title: Purification and Characterization of His-Tagged Recombinant Bacteroides fragilis Toxin-2 Variants In Vitro and In Vivo

doi: 10.3390/toxins18040189

Figure Lengend Snippet: Functional characterization of His-tagged BFT variants: ( a ) E-cadherin cleavage activity of A-hBFT from three His-rETBF clones. A-hBFT cleaves full-length E-cadherin (120 kDa) while serum-free medium (SFM) and imidazole controls do not. Clone 1 was selected for subsequent experiments. ( b ) Serial dilution analysis (1:10 1 to 1:10 8 ) comparing biological activity of His-tagged and wild-type BFT culture supernatants. ( c ) Cross-species validation in mouse (CMT93) and human (HT29/c1) epithelial cells. A-hBFT cleaves E-cadherin in both cell types while I-hBFT serves as negative control. Soluble 80 kDa E-cadherin ectodomain (sECAD) appears in culture supernatants following A-hBFT treatment. ( d ) RT-qPCR analysis of inflammatory cytokine response. A-hBFT significantly upregulates KC / cxcl1 (CMT93) and IL-8 / cxcl8 (HT29/c1) mRNA expression. Data represent mean ± SEM. ** p < 0.01, *** p < 0.001. ns = not significant.

Article Snippet: Human colonic epithelial cell line HT29/c1 (#HTB-38, ATCC, Manassas, VA, USA; passage 130) and mouse rectal epithelial cell line CMT93 (#CCL-223, ATCC, VA, USA; passage 75) were maintained in Dulbecco’s Modified Eagle Medium (DMEM, #MD10000A, Bandio Bio Science, Pocheon, Republic of Korea) supplemented with 5% fetal bovine serum (FBS, #35-015-CV, Corning, Palo Alto, CA, USA), 20 mM HEPES (#15630-080, Gibco, Miami, FL, USA), without penicillin-streptomycin [ , ].

Techniques: Functional Assay, Activity Assay, Clone Assay, Serial Dilution, Biomarker Discovery, Negative Control, Quantitative RT-PCR, Expressing

Journal: Toxins

Article Title: Purification and Characterization of His-Tagged Recombinant Bacteroides fragilis Toxin-2 Variants In Vitro and In Vivo

doi: 10.3390/toxins18040189

Figure Lengend Snippet: BFT receptor-dependent cellular binding patterns across mouse intestinal epithelial cell lines: Confocal microscopy of mouse intestinal epithelial cells treated with SFM, I-hBFT, or A-hBFT. Green fluorescence indicates hBFT detection via anti-6× His tag. ( a ) CMT93 cells (BFT-responsive) show binding only with A-hBFT treatment; ( b ) MSIE cells (BFT-non-responsive) show no binding under any condition; ( c ) YAMC cells (BFT-non-responsive) similarly show no binding signals. Differential binding patterns suggest BFT responsiveness depends on specific receptor presence rather than direct E-cadherin interaction. Scale bar = 20 μm.

Article Snippet: Human colonic epithelial cell line HT29/c1 (#HTB-38, ATCC, Manassas, VA, USA; passage 130) and mouse rectal epithelial cell line CMT93 (#CCL-223, ATCC, VA, USA; passage 75) were maintained in Dulbecco’s Modified Eagle Medium (DMEM, #MD10000A, Bandio Bio Science, Pocheon, Republic of Korea) supplemented with 5% fetal bovine serum (FBS, #35-015-CV, Corning, Palo Alto, CA, USA), 20 mM HEPES (#15630-080, Gibco, Miami, FL, USA), without penicillin-streptomycin [ , ].

Techniques: Binding Assay, Confocal Microscopy, Fluorescence

Journal:

Article Title: Stimulation of Chondrocyte Hypertrophy by Chemokine Stromal Cell-Derived Factor 1 in the Chondro-osseous Junction during Endochondral Bone Formation

doi: 10.1016/j.ydbio.2010.02.033

Figure Lengend Snippet: To determine whether SDF-1 can diffuse into cartilage, 17-day-chicken embryonic sternal cartilage was incubated with SDF-1 (100ng/mL) or without SDF-1 for 1h, 3h, and 24h. 10 µm frozen sections were used to detect SDF-1 by immuno-fluorescent staining with mAb against SDF-1. Fluorescence microscopy showed a progressive increase in SDF-1 staining (red color) surrounding chondrocytes during the 24 h time course (A, B,C) compared to control at 24 h (D). Scale bar = 20 µm.

Article Snippet: Before collecting samples for experiment, the cells were stimulated with SDF-1 (100ng/mL; Cat# 351-FS, R&D Systems, Inc. Minneapolis, MN) for 24h or pretreated with AMD3100 for 2 h (5ug/mL; Cat# 155148–31–5, Sigma-Aldrich, St. Louis, MO), a specific inhibitor for CXCR4, before stimulation with SDF-1.

Techniques: Incubation, Staining, Fluorescence, Microscopy, Control

Journal:

Article Title: Stimulation of Chondrocyte Hypertrophy by Chemokine Stromal Cell-Derived Factor 1 in the Chondro-osseous Junction during Endochondral Bone Formation

doi: 10.1016/j.ydbio.2010.02.033

Figure Lengend Snippet: To confirm that SDF-1 induces chondrocyte hypertrophy in growth plates, 12-day-old chicken tibia growth plates were cultured in the presence of SDF-1 (100ng/mL) or in the absence of SDF-1 for 2, 4, and 6 days. 10 µm frozen sections were used to detect Type X collagen expression by immuno-fluorescent staining with mAb against type X collagen. A progressive increase in the size of the hypertrohic growth plate based on Type X collagen staining was seen. The ratio of the length of the hypertrophic zone to that of the total growth plate was calculated at the different time points (B). (* p<0.05). Scale bar = 100 µm.

Article Snippet: Before collecting samples for experiment, the cells were stimulated with SDF-1 (100ng/mL; Cat# 351-FS, R&D Systems, Inc. Minneapolis, MN) for 24h or pretreated with AMD3100 for 2 h (5ug/mL; Cat# 155148–31–5, Sigma-Aldrich, St. Louis, MO), a specific inhibitor for CXCR4, before stimulation with SDF-1.

Techniques: Cell Culture, Expressing, Staining

Journal: Virologica Sinica

Article Title: Cytoplasmic domain and enzymatic activity of ACE2 are not required for PI4KB dependent endocytosis entry of SARS-CoV-2 into host cells

doi: 10.1016/j.virs.2022.03.003

Figure Lengend Snippet: PI4KB is relevant to S pseudovirus infection. A Diagram of phosphoinositide metabolic pathways. PIK93 and wortmannin were used as inhibitors targeting PI4K. B 293T-ACE2 cells were transfected with HA-Sac1 or HA-INPP5E before they were transduced with S pseudovirus and luciferase was measured. Values were normalized to control and expressed as mean ± SD. Inset, immunoblotting of HA-Sac1 or HA-INPP5E expression with anti-HA antibody. C Huh 7 cells pretreated with indicated concentrations of PIK93 or wortmannin were transduced with S pseudovirus and luciferase was measured. Values were normalized to control and expressed as mean ± SD. D Huh 7 cells transduced with the indicated shRNAs were infected with S pseudovirus and luciferase was measured to reflect viral infection (black bars). PI4KB mRNA was quantified by qRT-PCR (open bars). Values were normalized to control shRNA. PI4KB, phosphatidylinositol 4-kinase β; PI, Phosphoinositides; INPP5E, inositol polyphosphate-5-phosphatase E; PIP5K, Phosphatidylinositol-4-phosphate 5-kinase; S, spike; SD, standard deviation.

Article Snippet: PIK93 (#HY-12046), Wortmannin (#HY-10197), camostat mesylate (#HY-13512) and E-64d (#HY-100229) were purchased from MedChemExpress.

Techniques: Infection, Transfection, Transduction, Luciferase, Control, Western Blot, Expressing, Quantitative RT-PCR, shRNA, Standard Deviation

Journal: Theranostics

Article Title: Cerebellar Purkinje cell firing promotes conscious recovery from anesthesia state through coordinating neuronal communications with motor cortex

doi: 10.7150/thno.89592

Figure Lengend Snippet: Regulation of M1 cortical activity by 5Cb Purkinje cells via thalamus. (A) Schematic representation of the virus injection sites in M1, Po and 5Cb of Pcp2/L7-cre mice. (B) Viral injection sites for GCaMP in M1, and eGFP in Po and 5Cb. (C) Averaged CaMKIIα + neuron activity upon activation of Purkinje cells. Translucent red area indicates M1 CaMKIIα + neuron activity in mice treated with saline (left, ChR2+Saline); translucent blue area indicates M1 CaMKIIα + neuron activity in mice treated with CNO (right, ChR2+CNO). (D) Averaged ΔF/F values for ChR2+Saline and ChR2+CNO mice. (E) Schematic representation of the virus injection sites in M1, PF and 5Cb of Pcp2/L7-cre mice. (F-G) Viral injection sites for GCaMP in M1, and eGFP in PF and 5Cb. (H) Averaged CaMKIIα + neuron activity upon activation of Purkinje cells. Translucent red area indicates M1 CaMKIIα + neuron activity in ChR2+Saline mice; translucent blue area indicates M1 CaMKIIα+ neuron activity in ChR2+CNO mice. (I) Averaged ΔF/F values for ChR2+Saline and ChR2+CNO mice. Data are means ± SEM (n=5). M1: primary motor cortex; Po: posterior thalamic nuclear group; PF: parafascicular thalamic nucleus; 5Cb: the 5th lobule of the cerebellar vermis; CNO: Clozapine-N-oxide.

Article Snippet: Prior to the 30-minute photo-activation experiments, the mice received intraperitoneal injection of either saline or Clozapine-N-oxide (CNO, 2 mg/kg dissolved in saline; MedChemExpress).

Techniques: Activity Assay, Virus, Injection, Activation Assay, Saline

Journal: International Journal of Nanomedicine

Article Title: Integrating tacrolimus into eutectic oil-based microemulsion for atopic dermatitis: simultaneously enhancing percutaneous delivery and treatment efficacy with relieving side effects

doi: 10.2147/IJN.S212260

Figure Lengend Snippet: The AFM image of FK506 MCE ME. Abbreviations: AFM, atomic force microscope; FK506, tacrolimus; FK506 MCE ME, tacrolimus loaded microemulsion based on menthol/camphor eutectic.

Article Snippet: Abbreviations: AFM, atomic force microscope; FK506, tacrolimus; FK506 MCE ME, tacrolimus loaded microemulsion based on menthol/camphor eutectic.

Techniques: Microscopy

Journal: International Journal of Nanomedicine

Article Title: Integrating tacrolimus into eutectic oil-based microemulsion for atopic dermatitis: simultaneously enhancing percutaneous delivery and treatment efficacy with relieving side effects

doi: 10.2147/IJN.S212260

Figure Lengend Snippet: In vitro skin penetration of FK506 from different formulations ( A ) and skin retention of FK506 after 24 h penetration ( B ). Note: ** P <0.001. Abbreviations: FK506, tacrolimus; PC, The solution of 0.1% tacrolimus (FK506,w/v) dissolved in propylene carbonate; Oil free, 0.1% (w/v) FK506 solution containing the same surfactant and cosurfactant as microemulsion but without menthol/camphor eutectic; Ointment, 0.1%(w/w) FK506 commercial ointment; MCE ME, 0.1% (w/v) FK506 microemulsion based on menthol/camphor eutectic; MCE ME gel, 0.1% (w/w) FK506 microemulsion gel based on menthol/camphor eutectic; h, hours; SD, standard deviation.

Article Snippet: Abbreviations: AFM, atomic force microscope; FK506, tacrolimus; FK506 MCE ME, tacrolimus loaded microemulsion based on menthol/camphor eutectic.

Techniques: In Vitro, Ointment, Standard Deviation

Journal: International Journal of Nanomedicine

Article Title: Integrating tacrolimus into eutectic oil-based microemulsion for atopic dermatitis: simultaneously enhancing percutaneous delivery and treatment efficacy with relieving side effects

doi: 10.2147/IJN.S212260

Figure Lengend Snippet: Cell viability of HaCaT cells after incubation with MCE ME vehicle of different concentrations for 12 h and 24 h, respectively. Abbreviations: HaCaT, immortal human keratinocytes; MCE, ME vehicle; Microemulsion based on menthol/camphor eutectic without FK506; SD, standard deviation.

Article Snippet: Abbreviations: AFM, atomic force microscope; FK506, tacrolimus; FK506 MCE ME, tacrolimus loaded microemulsion based on menthol/camphor eutectic.

Techniques: Incubation, Standard Deviation

Journal: International Journal of Nanomedicine

Article Title: Integrating tacrolimus into eutectic oil-based microemulsion for atopic dermatitis: simultaneously enhancing percutaneous delivery and treatment efficacy with relieving side effects

doi: 10.2147/IJN.S212260

Figure Lengend Snippet: Schematic diagram of the experimental procedure ( A ); effects of different formulations containing 0.1% FK506 on dermatitis scores throughout the experiment ( B ); and clinical features of AD-like skin lesions in the end of experiment ( C ). Notes: * P <0.05 in comparison with DNCB; ** P <0.001 in comparison with DNCB; # P <0.05 in comparison with DNCB + Ointment; ## P <0.001 in comparison with DNCB + Ointment; ++ P <0.001. Abbreviations: AD, atopic dermatitis; DNCB, 1–chloro–2,4–dinitrobenzene; FK506, tacrolimus; Control, healthy mice with shaved dorsal region; DNCB, AD-induced mice without drug treatment; DNCB + PC, AD-induced mice treated with propylene carbonate containing 0.1% (w/v) FK506; DNCB + Ointment; AD-induced mice treated with commercial ointment containing 0.1% (w/w) FK506; DNCB + Oil free, AD-induced mice treated with Oil free containing 0.1% (w/v) FK506; DNCB + Vehicle, AD-induced mice treated with microemulsion based on menthol/camphor eutectic vehicle without FK506; DNCB + MCE ME, AD-induced mice treated with microemulsion based on menthol/camphor eutectic containing 0.1% (w/v) FK506; DNCB + MCE ME gel, AD-induced mice treated with microemulsion based on menthol/camphor eutectic gel containing 0.1% (w/w) FK506; SD, standard deviation.

Article Snippet: Abbreviations: AFM, atomic force microscope; FK506, tacrolimus; FK506 MCE ME, tacrolimus loaded microemulsion based on menthol/camphor eutectic.

Techniques: Comparison, Ointment, Control, Standard Deviation

Journal: International Journal of Nanomedicine

Article Title: Integrating tacrolimus into eutectic oil-based microemulsion for atopic dermatitis: simultaneously enhancing percutaneous delivery and treatment efficacy with relieving side effects

doi: 10.2147/IJN.S212260

Figure Lengend Snippet: The ear thickness ( A ), spleen index ( B ) on day 28, and TEWL ( C ) of the mice treated with different formulations throughout the experiment. Notes: * P <0.05 in comparison with DNCB; ** P <0.001 in comparison with DNCB; # P <0.05 in comparison with DNCB + Ointment; ## P <0.001 in comparison with DNCB +Ointment; + P <0.05; ++ P <0.001. Abbreviations: AD, atopic dermatitis; DNCB, 1–chloro–2,4–dinitrobenzene; FK506, tacrolimus; TEWL, transdermal water loss; Control, healthy mice with shaved dorsal region; DNCB, AD-induced mice without drug treatment; DNCB + PC, AD-induced mice treated with propylene carbonate containing 0.1% (w/v) FK506; DNCB + Ointment: AD-induced mice treated with commercial ointment containing 0.1% (w/w) FK506; DNCB + Oil free, AD-induced mice treated with Oil free containing 0.1% (w/v) FK506; DNCB + Vehicle; AD-induced mice treated with microemulsion based on menthol/camphor eutectic vehicle without FK506; DNCB + MCE ME, AD-induced mice treated with microemulsion based on menthol/camphor eutectic containing 0.1% (w/v) FK506; DNCB + MCE ME gel, AD-induced mice treated with microemulsion based on menthol/camphor eutectic gel containing 0.1% (w/w) FK506; SD, standard deviation.

Article Snippet: Abbreviations: AFM, atomic force microscope; FK506, tacrolimus; FK506 MCE ME, tacrolimus loaded microemulsion based on menthol/camphor eutectic.

Techniques: Comparison, Ointment, Control, Standard Deviation

Journal: International Journal of Nanomedicine

Article Title: Integrating tacrolimus into eutectic oil-based microemulsion for atopic dermatitis: simultaneously enhancing percutaneous delivery and treatment efficacy with relieving side effects

doi: 10.2147/IJN.S212260

Figure Lengend Snippet: Histological features of the skin treated with different formulations in AD mouse model. The sections were stained with hematoxylin and eosin ( A ) and toluidine blue ( B ), respectively. Abbreviations: AD, atopic dermatitis; DNCB, 1–chloro–2,4–dinitrobenzene; FK506, tacrolimus; Control, healthy mice with shaved dorsal region; DNCB, AD-induced mice without drug treatment; DNCB + PC, AD-induced mice treated with propylene carbonate containing 0.1% (w/v) FK506; DNCB + Ointment; AD-induced mice treated with commercial ointment containing 0.1% (w/w) FK506; DNCB + Oil free; AD-induced mice treated with Oil free containing 0.1% (w/v) FK506; DNCB + Vehicle, AD-induced mice treated with microemulsion based on menthol/camphor eutectic vehicle without FK506; DNCB + MCE ME, AD-induced mice treated with microemulsion based on menthol/camphor eutectic containing 0.1% (w/v) FK506; DNCB + MCE ME gel, AD-induced mice treated with microemulsion based on menthol/camphor eutectic gel containing 0.1% (w/w) FK506.

Article Snippet: Abbreviations: AFM, atomic force microscope; FK506, tacrolimus; FK506 MCE ME, tacrolimus loaded microemulsion based on menthol/camphor eutectic.

Techniques: Staining, Control, Ointment

Journal: International Journal of Nanomedicine

Article Title: Integrating tacrolimus into eutectic oil-based microemulsion for atopic dermatitis: simultaneously enhancing percutaneous delivery and treatment efficacy with relieving side effects

doi: 10.2147/IJN.S212260

Figure Lengend Snippet: Effects of different formulations on serum total IgE. Notes: * P <0.05 in comparison with DNCB; ** P <0.001 in comparison with DNCB; # P <0.05 in comparison with DNCB + Ointment; ## P <0.001 in comparison with DNCB +Ointment; + P <0.05; ++ P <0.001. Abbreviations: AD, atopic dermatitis; DNCB, 1–chloro–2,4–dinitrobenzene; FK506, tacrolimus; Control, healthy mice with shaved dorsal region; DNCB, AD-induced mice without drug treatment; DNCB + PC, AD-induced mice treated with propylene carbonate containing 0.1% (w/v) FK506; DNCB + Ointment: AD-induced mice treated with commercial ointment containing 0.1% (w/w) FK506; DNCB + Oil free, AD-induced mice treated with Oil free containing 0.1% (w/v) FK506; DNCB + Vehicle, AD-induced mice treated with microemulsion based on menthol/camphor eutectic vehicle without FK506; DNCB + MCE ME, AD-induced mice treated with microemulsion based on menthol/camphor eutectic containing 0.1% (w/v) FK506; DNCB + MCE ME gel, AD-induced mice treated with microemulsion based on menthol/camphor eutectic gel containing 0.1% (w/w) FK506; SD, standard deviation.

Article Snippet: Abbreviations: AFM, atomic force microscope; FK506, tacrolimus; FK506 MCE ME, tacrolimus loaded microemulsion based on menthol/camphor eutectic.

Techniques: Comparison, Ointment, Control, Standard Deviation

Journal: International Journal of Nanomedicine

Article Title: Integrating tacrolimus into eutectic oil-based microemulsion for atopic dermatitis: simultaneously enhancing percutaneous delivery and treatment efficacy with relieving side effects

doi: 10.2147/IJN.S212260

Figure Lengend Snippet: Inhibition effects of different formulations on scratch frequency ( A ) and substance P expression ( B ). Notes: * P <0.05 in comparison with DNCB; ** P <0.001 in comparison with DNCB; # P <0.05 in comparison with DNCB + Ointment; ## P <0.001 in comparison with DNCB +Ointment; + P <0.05; ++ P <0.001. Abbreviations: AD, atopic dermatitis; DNCB, 1–chloro–2,4–dinitrobenzene; FK506, tacrolimus; Control, healthy mice with shaved dorsal region; DNCB, AD-induced mice without drug treatment; DNCB + PC, AD-induced mice treated with propylene carbonate containing 0.1% (w/v) FK506; DNCB + Ointment: AD-induced mice treated with commercial ointment containing 0.1% (w/w) FK506; DNCB + Oil free, AD-induced mice treated with Oil free containing 0.1% (w/v) FK506; DNCB + Vehicle, AD-induced mice treated with microemulsion based on menthol/camphor eutectic vehicle without FK506; DNCB + MCE ME, AD-induced mice treated with microemulsion based on menthol/camphor eutectic containing 0.1% (w/v) FK506; DNCB + MCE ME gel, AD-induced mice treated with microemulsion based on menthol/camphor eutectic gel containing 0.1% (w/w) FK506; SD, standard deviation.

Article Snippet: Abbreviations: AFM, atomic force microscope; FK506, tacrolimus; FK506 MCE ME, tacrolimus loaded microemulsion based on menthol/camphor eutectic.

Techniques: Inhibition, Expressing, Comparison, Ointment, Control, Standard Deviation