Journal: International Dental Journal

Article Title: 4632427E13Rik Facilitates Jaw Marrow-Derived Mesenchymal Stem Cells Osteogenesis and Angiogenesis Under Hypoxia Through miR-34a-5p/Aldoa/Hif-1α Pathway

doi: 10.1016/j.identj.2025.109364

Figure Lengend Snippet: Aldoa promotes osteogenesis and angiogenesis via the ERK/Hif-1α pathway. A, Alizarin red staining (14 days) and ALP staining (7 days). B, The gene expression levels of Runx2, Alp and Ocn were determined by qRT-PCR. β-actin was used as an internal reference gene. C, Tube formation assay was performed in the presence of CM. The scale bars represent 100 μm. D, The mRNA expression levels of Vegf and CD31 were analysed by quantitative RT-PCR. β-actin was used as an internal reference gene. E, Immunofluorescence staining was performed to detect Vegf expression after being treated by CM. The scale bars represent 25 μm. F, A Kyoto Encyclopaedia of Genes and Genomes (KEGG) enrichment analysis was performed to determine the top related pathways involving these differentially expressed mRNAs. G and H, The protein expression levels of Aldoa, Hif-1α and the phosphorylated of ERK, JUK and p38 among hypoxia, normoxia and hypoxia + si-Aldoa groups were determined by western blot. Semiquantitative analysis of the p-ERK/ERK, p-JUK/JUK and p38/p-p38 ratios were shown. β-actin was used as an internal reference gene. All data were expressed as means ± SD. * P < .05, ** P < .01.

Article Snippet: Rabbit anti-Aldoa (1:1000, 11217-1-AP), rabbit anti-Hif-1α (1:1000, ab179483), rabbit anti-Runx2 (1:1000, 8486S), rabbit anti-CD31 (1:1000, 77699S), rabbit anti-Ocn (1:1000, bs-4917R), rabbit anti-Alp (1:1000, bs-1535R), rabbit anti-Vegf (1:1000, bs-1313R), rabbit anti-ERK (1:1000, bsm-33337M), rabbit anti-p-ERK (1:1000, bs-1646R), rabbit anti-p38 (1:1000, bs-0637R), rabbit anti-p-p38 (1:1000, bs-0636R), rabbit anti-JNK (1:1000, bs-2592R), rabbit anti-p-JNK (1:1000, bs-1640R), and mouse anti-β-actin (1:2000, AF7018) were purchased from Proteintech, Abcam, Cell Signalling Technology, and Bioss, respectively.

Techniques: Staining, Gene Expression, Quantitative RT-PCR, Tube Formation Assay, Expressing, Immunofluorescence, Western Blot

Journal: International Dental Journal

Article Title: 4632427E13Rik Facilitates Jaw Marrow-Derived Mesenchymal Stem Cells Osteogenesis and Angiogenesis Under Hypoxia Through miR-34a-5p/Aldoa/Hif-1α Pathway

doi: 10.1016/j.identj.2025.109364

Figure Lengend Snippet: Aldoa promotes osteogenesis and angiogenesis via the ERK/Hif-1α pathway. A, Alizarin red staining (14 days) and ALP staining (7 days). B, The gene expression levels of Runx2, Alp and Ocn were determined by qRT-PCR. β-actin was used as an internal reference gene. C, Tube formation assay was performed in the presence of CM. The scale bars represent 100 μm. D, The mRNA expression levels of Vegf and CD31 were analysed by quantitative RT-PCR. β-actin was used as an internal reference gene. E, Immunofluorescence staining was performed to detect Vegf expression after being treated by CM. The scale bars represent 25 μm. F, A Kyoto Encyclopaedia of Genes and Genomes (KEGG) enrichment analysis was performed to determine the top related pathways involving these differentially expressed mRNAs. G and H, The protein expression levels of Aldoa, Hif-1α and the phosphorylated of ERK, JUK and p38 among hypoxia, normoxia and hypoxia + si-Aldoa groups were determined by western blot. Semiquantitative analysis of the p-ERK/ERK, p-JUK/JUK and p38/p-p38 ratios were shown. β-actin was used as an internal reference gene. All data were expressed as means ± SD. * P < .05, ** P < .01.

Article Snippet: Rabbit anti-Aldoa (1:1000, 11217-1-AP), rabbit anti-Hif-1α (1:1000, ab179483), rabbit anti-Runx2 (1:1000, 8486S), rabbit anti-CD31 (1:1000, 77699S), rabbit anti-Ocn (1:1000, bs-4917R), rabbit anti-Alp (1:1000, bs-1535R), rabbit anti-Vegf (1:1000, bs-1313R), rabbit anti-ERK (1:1000, bsm-33337M), rabbit anti-p-ERK (1:1000, bs-1646R), rabbit anti-p38 (1:1000, bs-0637R), rabbit anti-p-p38 (1:1000, bs-0636R), rabbit anti-JNK (1:1000, bs-2592R), rabbit anti-p-JNK (1:1000, bs-1640R), and mouse anti-β-actin (1:2000, AF7018) were purchased from Proteintech, Abcam, Cell Signalling Technology, and Bioss, respectively.

Techniques: Staining, Gene Expression, Quantitative RT-PCR, Tube Formation Assay, Expressing, Immunofluorescence, Western Blot

Journal: bioRxiv

Article Title: An APP-centered molecular gateway integrates innate immunity and retinoic acid signaling to drive irreversible metamorphic commitment

doi: 10.64898/2026.01.22.700939

Figure Lengend Snippet: (A) Settlement rate of inhibitor-treated larvae. The box plots with superimposed jitter plots display the larval settlement rate under various inhibitor treatments. The biofilm stimulus condition is used as the positive control. The concentration of each inhibitor is indicated on the horizontal axis. The data represent the settlement rate of larvae remaining attached out of 10 larvae across six independent biological replicates (total n = 60). Statistical significance among treatment groups was assessed using one-way ANOVA followed by Tukey’s HSD post hoc test, with grouping letters indicating significant differences ( p < 0.05); treatments sharing a letter are not significantly different. (B) Quantitative assessment of the functional hierarchy. The box plots with superimposed jitter plots show the Metamorphic Progression Scores (MPS) for larvae treated with various pharmacological inhibitors with or without all-trans retinoic acid (RA). The MPS was calculated based on the metamorphic stage reached by the larvae in the identical assays used for the settlement rate analysis in (A). The concentration of each inhibitor is indicated on the horizontal axis. The MPS represents the average metamorphic stage reached (0 = brachiolaria; 1 = early; 2 = middle; 3 = late; 4 = pre-juvenile; 5 = juvenile). Statistical significance among the treatment groups was assessed using one-way ANOVA followed by Tukey’s HSD post hoc test ( *p < 0.05; n.s., not significant). A significant RA-dependent rescue condition (a statistically significant increase in MPS compared with the inhibitor-alone condition) is highlighted in grey, establishing the functional hierarchy of the pathways relative to the RA commitment signal. (C) Representative image illustrating pathway functional hierarchy. Images show representative larval morphology under the control, inhibitor-only, and inhibitor + RA conditions. These images specifically represent the high-concentration inhibitor treatments (MyD88 inhibitor: 50 µM; MAPK inhibitors: 10 µM; IKKβ and HSP90AA1 inhibitors: 1 µM). MyD88 inhibition completely blocks the behavioral decision of settlement. JNK and p38 inhibition caused a distinct early-stage arrest (low MPS), and the effects of their inhibition were significantly rescued by RA co-treatment. In contrast, ERK inhibition arrested metamorphosis at the middle stage, and this block was not rescued by exogenous RA. Similarly, IKKβ and HSP90AA1 inhibition arrested metamorphosis at later stages, and this block was not rescued by exogenous RA, functionally placing all three pathways (ERK, IKKβ, and HSP90AA1) downstream of the RA commitment signal. Scale bar: 200 µm. Inhibitors used: T6167923 (MyD88 inhibitor), IKK-16 (IKKβ inhibitor), U0126 (ERK inhibitor), SP600125 (JNK inhibitor), SB202190 (p38 inhibitor), and Luminespib (HSP90AA1 inhibitor).

Article Snippet: The inhibitors were dissolved in DMSO and applied at the indicated concentrations: the MyD88 inhibitor T6167923 (5 or 50 μM; MedChemExpress), the IKKβ inhibitor IKK-16 (0.1 or 1 μM; MedChemExpress), and MAPK inhibitors SP600125 (JNK), SB202190 (p38), and U0126 (ERK) (1 or 10 μM; MedChemExpress or FUJIFILM Wako Pure Chemical Corporation), and the HSP90AA1 inhibitors luminespib (0.1 or 1 μM; Chemscene).

Techniques: Positive Control, Concentration Assay, Functional Assay, Control, Inhibition, Blocking Assay

Journal: bioRxiv

Article Title: An APP-centered molecular gateway integrates innate immunity and retinoic acid signaling to drive irreversible metamorphic commitment

doi: 10.64898/2026.01.22.700939

Figure Lengend Snippet: This model illustrates the proposed three-tiered molecular switch that translates external microbial cues into the irreversible developmental fate of sea star metamorphosis, based on Dynamic Network Module (DNM) analysis and comprehensive pharmacological functional assays. This cascade integrates innate immune and developmental signaling pathways across three functional layers: Signal Sensing, Commitment Conversion, and Irreversible Execution. The process is initiated in the Signal Sensing layer, where the environmental cue, microbial biofilms, activates the adapter protein MyD88, which serves as an obligatory first-tier hub. MyD88 transmits signals via the JNK/p38/ERK MAPK pathway to govern the initial settlement behavior. MyD88 exhibits a concentration-dependent dual output: high-dose inhibition abolishes settlement behavior (RA-non-rescuable), while low-dose inhibition permits settlement but causes a late-stage molecular arrest (RA-non-rescuable). Following sensing, the cascade enters the Commitment Conversion layer. JNK/p38 MAPK acts as an essential hybrid adapter that converts immune signals into a Retinoic Acid (RA) hormonal commitment signal (RA-rescuable phenotype). The Amyloid Precursor Protein (APP) functions as the irrevocable commitment gateway, integrating inputs from the upstream MAPK, IKKβ/NFκB, and RA signaling axes to make the final molecular decision. APP ensures irreversibility through “signal focusing,” maintaining its signal strength during the systemic “mass shutdown” of non-essential larval programs. The process culminates in an Irreversible Execution Tier, where the robust execution of the metamorphic program relies on the multi-layered convergence of signals onto the master transcription factor, TFAP2A. The APP commitment decision is translated into transcriptional output via the release of its intracellular domain (AICD), which acts as the final dedicated execution switch by converging to TFAP2A in complex with GSK3β/Src. TFAP2A receives parallel inputs from RA (for launching the program), IKKβ/NFκB (for sustained maintenance and transcriptional output; RA non-rescuable), and ERK (a crucial early execution factor for immediate morphogenesis and physical attachment maintenance; RA non-rescuable). Finally, the RA signal induces the HSP90AA1 chaperone, establishing a positive feedback loop that maintains the structural integrity and function of critical signaling complexes (including MyD88 and APP), thereby ensuring the stability of the executed program.

Article Snippet: The inhibitors were dissolved in DMSO and applied at the indicated concentrations: the MyD88 inhibitor T6167923 (5 or 50 μM; MedChemExpress), the IKKβ inhibitor IKK-16 (0.1 or 1 μM; MedChemExpress), and MAPK inhibitors SP600125 (JNK), SB202190 (p38), and U0126 (ERK) (1 or 10 μM; MedChemExpress or FUJIFILM Wako Pure Chemical Corporation), and the HSP90AA1 inhibitors luminespib (0.1 or 1 μM; Chemscene).

Techniques: Functional Assay, Protein-Protein interactions, Concentration Assay, Inhibition