Jing Meng

Background: ASARI RADIX ET RHIZOMA (Xixin), extensively utilized in clinical settings, contains aristolochic acid analogs (AAAs), primarily AA-IVa, with minor quantities of AL-I and negligible levels of AA-I. AA-I mediates aristolochic acid-induced nephropathy; therefore, the safety of Xixin is controversial. The toxicity of AAAs is varied, and the mechanisms are ambiguous. Objective: To compare the toxicity of AA-I, AA-IVa, and AL-I toward renal tubular epithelial cells and explain the molecular causes of such. Materials and Methods: C57BL/6 mice were administered AA-I, AA-IVa, or AL-I for 24 weeks; renal toxicity was assessed via survival and histopathology. In vitro, AAAs toxicity was evaluated in human renal tubular epithelial cells (HK-2) by high-content Hoechst 33342/PI apoptotic tests and fibrosis marker quantification. Transcriptomics revealed alterations in genes and pathways induced by AAAs. Click chemistry-based activity-based protein profiling, molecular docking, surface plasmon resonance, enzyme activity assays, and gene knockdown identified the AAAs’ direct targets. Results: AA-I induced dose-dependent mortality, severe kidney injury, and fibrosis; AL-I showed less toxicity, and AA-IVa was not discernibly toxic in mice. In vitro, AA-I prompted apoptosis and fibrosis in HK-2 via transcriptional modification; other AAAs exhibited diminished effects. AA-I, AL-I and AA-IVa showed stable binding ability to HDAC2; AA-I and AL-I enhanced HDAC2 expression; and AA-I facilitated HDAC2 nuclear translocation and enzymatic activity. AA-I, unlike AL-I and AA-IVa, augmented MYC expression, disturbed the BCL2/BAX equilibrium, activated Caspase-7, and triggered apoptosis and pro-fibrotic transformation. Conclusions: AA-I induces apoptosis and fibrosis through HDAC2, and HDAC2 partially contributes to the various toxicities of AA-I, AA-IVa, and AL-I. This study provides fundamental evidence for the safety evaluation of Xixin.

Background: Dangguibuxue decoction (DD), containing Angelica sinensis (Oliv.) Diels (AS) and Astragalus membranaceus (Fisch.) Bge. (AM) (1:5), is a well-known traditional Chinese medicine (TCM) used for strengthening qi and nourishing the blood. DD has shown therapeutic effects in nephropathy patients. However, the underlying mechanisms based on the traditional efficacy are still not fully elucidated. Methods: The chemical constituents in DD were identified using UPLC-MS/MS. Network pharmacology analysis was applied to predict the potential target genes and associated signaling pathways. A renal fibrosis mouse model was induced by the intraperitoneal injection of aristolochic acid I (AA I) at 3.0 mg/kg. Mice were treated with AM, AS, and DD at two dosages by oral gavage for 30 days. Body weights, serum biochemistry, hematology, and histopathology observations were assessed. The key targets predicted were validated using qRT-PCR and Western blotting. The active constituents were screened by molecular docking, and their anti-fibrotic effects were evaluated through in vitro assays. Results: DD effectively improved renal functions and alleviated AA I-induced renal fibrosis. DD alleviated anemia and upregulated the expression of Erythropoietin (EPO). Network pharmacology analysis indicated the involvement of signaling pathways, including the PI3K/Akt, hypoxia-inducible factor-1α (HIF-1α) and transforming growth factor-β (TGF-β) signaling pathways. Experimental validation further demonstrated that DD reduced the protein expression of HIF-1α, collagen I, and TGF-β, and the ratios of phosphorylated Smad2/3 to total Smad2/3. Molecular docking and in vitro assays suggested that rutin may be a potential bioactive compound in DD. Conclusions: This research indicated that DD ameliorated AA I-induced renal fibrosis in mice, which may be associated with the modulation of HIF-1α and TGF-β/Smad signaling pathways. Rutin may be a potential bioactive compound in DD with anti-fibrotic activity, but further studies are still needed to clarify the content of rutin in DD, the amount of its exposure in the body, and its contribution to the effects of DD.

Triptonide (TN), a natural bioactive compound derived from Tripterygium wilfordii with multiple antitumor activities, has a poorly defined exact mechanism in acute myeloid leukemia (AML)—a hematologic malignancy with limited treatment options. This study systematically clarifies TN’s mechanisms in AML through an integrative strategy combining network pharmacology, molecular docking, molecular dynamics simulation, and in vitro/in vivo experiment validation. Predicted TN targets using Swiss Target Prediction and PharmMapper, and AML-associated genes via GeneCards, OMIM, and CTD. Verall, O198 overlapping targets were mapped to build a PPI network using STRING and Cytoscape. Identified hub gene (AKT1, EGFR, HSP90AA1, HSP90AB1, and PIK3R1) using CytoNCA, MCODE, and CytoHubba algorithms. GO and KEGG enrichment analyses highlighted marked enrichment in the PI3K/AKT pathway. TN exhibited high affinity binding to AKT1 (−7.28 kcal/mol) and PIK3R1 (−7.33 kcal/mol), with stable interactions confirmed by molecular dynamics simulations. The GSEA of the DEGs from GEPIA 2 revealed prominent activation of the PI3K/AKT signaling pathway, indicating its key role as a regulator of AML pathogenesis. In vitro, TN dose-dependently suppressed proliferation of multiple AML cell lines induced apoptosis, and downregulated the expression of P-PI3K and P-AKT. The AKT activator SC79 reversed TN-induced suppression in AML cells, validating PI3K/AKT pathway dependency. In vivo, TN significantly inhibited tumor growth in xenograft models without causing organ toxicity in female nude mice. These findings reveal the therapeutic potential of TN against AML through inhibiting the PI3K/AKT axis. With no PI3K/AKT inhibitors targeting AML approved as first-line therapies, TN emerges as a promising candidate for AML treatment, offering a safer natural alternative.

Objective: To investigate electroacupuncture (EA)-induced reorganization of voxel-mirrored homotopic connectivity (VMHC) and its association with motor recovery in a rat model of ischemic stroke. Methods: Twenty-four female Sprague–Dawley rats were randomized into Sham group, stroke model group and EA group. The EA group received 1-week treatment (2/15 Hz sparse-dense wave, 2 mA, 30 min/day) at GV20. Neurological deficits were assessed using the modified neurological severity score. And resting-state functional magnetic resonance imaging was acquired pre-post-intervention for VMHC analysis. Group differences and VMHC-behavior correlations were evaluated. Results: EA significantly increased VMHC in subcortical motor regions ( p = 0.001, alphasim correction) but decreased VMHC in somatosensory cortex versus untreated stroke. Model group showed progressive VMHC reductions in hippocampus, hypothalamus, and somatosensory cortex. Enhanced internal capsule VMHC correlated with improved Longa scores in EA rats ( r ² = 0.206, p = 0.009). Conclusion: EA promotes motor recovery through frequency-specific bidirectional VMHC modulation. This study elucidates EA’s inter-hemispheric connectivity level therapeutic mechanism for stroke rehabilitation.