1.Mechanisms of Yishen Juanbi Pills-containing Bone Marrow Fluid in Improving Kidney-deficiency Rheumatoid Arthritis by Regulating CD4+ T Lymphocyte Differentiation via SDF-1/CXCR4 Signaling Pathway
Jinlin TONG ; Yuyao WANG ; Hong LIU ; Jinghua PAN ; Danping FAN ; Hongyan ZHAO
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(4):90-99
ObjectiveTo investigate the effects of Yishen Juanbi pills (YSJB)-containing bone marrow fluid on the migration and differentiation phenotypes of CD4⁺T lymphocytes based on the stromal cell-derived factor-1/chemokine receptor 4 (SDF-1/CXCR4) signaling pathway. MethodsPrimary CD4⁺T lymphocytes were isolated from mice using magnetic bead separation and identified for purity by flow cytometry. A CD4⁺T lymphocyte culture system was then established to observe the effects of SDF-1 on CD4⁺T-cell migration and differentiation. On this basis, the experimental groups included the Sham group, the ovariectomy (OVX) group, the Sham+collagen-induced arthritis (CIA) group, the OVX+CIA group, the Sham+CIA+YSJB group (2.16 g·kg-1), the OVX+CIA+YSJB group (2.16 g·kg-1), and the OVX+CIA+methotrexate (MTX) group (1.5 mg·kg-1). Bone marrow fluid from each group was prepared according to previous methods and added to the CD4⁺ T-cell culture system at 5% (v/v). Transwell assays were used to examine CD4⁺T-cell migration in each group. Real-time PCR was used to measure the mRNA expression levels of interleukin (IL)-17, tumor necrosis factor-α (TNF-α), retinoic-acid-related orphan receptor γt (RORγt), IL-10, transforming growth factor-β (TGF-β), forkhead box P3 (FoxP3), CXCR4, phosphoinositide 3-kinase (PI3K), and protein kinase B (Akt). Western blot was used to detect the expression of helper T (Th)17/regulatory T (Treg) cell signature factors (RORγt, FoxP3), CXCR4, PI3K, phosphorylated (p)-PI3K, Akt, and p-Akt. In a separate set of experiments, cells were divided into the Sham group, OVX+CIA group, OVX+CIA+CXCR4 antagonist AMD3100 group, and OVX+CIA+YSJB+AMD3100 group to observe changes in the above indicators following AMD3100 intervention. ResultsCompared with the Sham group, the number of migrated cells in the lower chamber was significantly increased in the Sham+CIA and OVX+CIA groups (P<0.05, P<0.01). The mRNA expression of RORγt, IL-17, TNF-α, CXCR4, PI3K, and Akt was significantly upregulated, whereas FoxP3, IL-10, and TGF-β mRNA expression was significantly decreased (P<0.05, P<0.01). Protein expression of RORγt, CXCR4, p-PI3K/PI3K, and p-Akt/Akt was significantly increased, while FoxP3 protein expression was markedly decreased (P<0.05, P<0.01). Compared with the OVX+CIA group, the OVX+CIA+YSJB group and OVX+CIA+MTX group showed significantly reduced migration (P<0.05), mRNA expression of RORγt, IL-17, TNF-α, CXCR4, PI3K, and Akt was also significantly decreased, while FoxP3, IL-10, and TGF-β mRNA expression was significantly increased (P<0.05, P<0.01). RORγt protein expression was significantly downregulated, and FoxP3 protein expression markedly upregulated (P<0.05). In the OVX+CIA+YSJB group, CXCR4, p-PI3K/PI3K, and p-Akt/Akt protein expression was significantly decreased (P<0.05). Compared with the OVX+CIA group, RORγt, CXCR4, PI3K, and Akt mRNA expression in CD4⁺T cells was significantly decreased in the OVX+CIA+AMD3100 group and the OVX+CIA+YSJB+AMD3100 group, while FoxP3 mRNA and protein expression was significantly upregulated (P<0.05, P<0.01). RORγt, CXCR4, p-PI3K/PI3K, and p-Akt/Akt protein expression was also markedly decreased (P<0.05, P<0.01). Compared with the OVX+CIA+AMD3100 group, the OVX+CIA+YSJB+AMD3100 group showed significantly decreased RORγt and Akt mRNA expression (P<0.05) and significantly lower p-Akt/Akt protein expression (P<0.05). ConclusionYSJB-containing bone marrow fluid suppresses CD4⁺T-cell migration and regulates Th17/Treg balance by downregulating Th17-associated signature factors and upregulating Treg-associated signature factors through inhibition of the SDF-1/CXCR4 signaling pathway and PI3K/Akt signaling pathway. The SDF-1/CXCR4 signaling pathway is one of the targets through which YSJB inhibits CD4⁺T-cell differentiation.
2.Reflections on Status Quo and Development Pathways of Traditional Chinese Medicine Technology Transfer in Context of Digital-intelligent Transformation
Jie ZHANG ; Jing XU ; Guangwei ZHENG ; Huayu ZHANG ; Chang LIU ; Xiaoxiao WEN ; Xishui PAN ; Bin WANG
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(4):235-240
As a distinctive resource of Chinese civilization, traditional Chinese medicine (TCM) technology transfer faces significant opportunities under the background of digital and intelligent transformation, while also being constrained by unique challenges such as the complexity of its theoretical system, lengthy industrial chains, and multidimensional policy restrictions, resulting in a "high-value-high-threshold" paradox. At present, TCM technology transfer is deeply trapped in a "threefold reluctance" dilemma, i.e., unwillingness to transfer, inability to transfer, and lack of capacity to transfer. Specifically, the disconnection between scientific research evaluation systems and market demand leads to low conversion rates of research achievements, unclear ownership and compliance risks suppress innovation incentives, and the absence of professional services intensifies supply-demand mismatches. This article systematically analyzes the specific characteristics of TCM technology transfer and proposes a breakthrough pathway centered on full-chain digital and intelligent transformation. By integrating technologies such as intelligent sorting systems, blockchain-based traceability, and AI diagnostic models, the TCM ecosystem spanning "cultivation-production-service" can be reconstructed. In terms of standardization, promoting the progression from "experience-based data conversion" to "data standardization" and further to "intelligent standardization" is advocated to resolve quality control challenges. For example, a "three-no-one-full" certification system can strengthen quality trust. Policy coordination should focus on optimizing mechanisms for the transformation of scientific and technological achievements, while exploring intellectual property securitization and risk-sharing models to stimulate research momentum. In terms of internationalization, reliance on the Belt and Road Initiative platform to promote the export of geo-authentic medicinal material brands and standards is recommended to build a dual-driven model of "technology plus culture". Looking ahead, through the construction of national-level databases, the cultivation of interdisciplinary talent, and the mutual recognition of international standards, a new paradigm of "scientific intelligent manufacturing" can be formed, providing systematic solutions for the modernization of TCM and global health governance.
3.Expert Consensus on Neurocritical Care Monitoring and Management in Beijing and Tibet(2025)
Drolma PHURBU ; Wenjin CHEN ; Heng ZHANG ; Jian ZHANG ; Xiaomeng WANG ; Guoying LIN ; Wenjun PAN ; Xiying GUI ; Xin CAI ; Chodron TENZIN ; Jianlei FU ; Qianwei LI ; TSEYANG ; Yijun LIU ; Bo LIU ; Tsering DROLMA ; Yudron SONAM ; KYILV ; Samdrup TSERING ; Wa DA ; Juan GUO ; Cheng QIU ; Huan CHEN ; Xiaoting WANG ; Yangong CHAO ; Dawei LIU ; Wenzhao CHAI ; Chenggong HU ; Wanhong YIN ; Shihong ZHU
Medical Journal of Peking Union Medical College Hospital 2026;17(1):59-72
Neurocritical care involves complex pathophysiological mechanisms, and its incidence is higher, injuries are more severe, and treatment is more challenging in high-altitude environments. This consensus, based on the latest domestic and international evidence-based medical data, establishes a standardized, goal-oriented framework for neurocritical care management applicable in high-altitude regions and nationwide. The consensus was developed following international standards for evidence quality assessment and underwent two rounds of Delphi expert consultation, resulting in 32 recommendation statements covering three parts: management systems, monitoring and assessment, and core strategies. Key updates include: advocating for the establishment of independent neurocritical care units and implementing precise tiered diagnosis and treatment based on the "Five Differences in Critical Care" concept; constructing a "trinity" multimodal brain monitoring system centered on cerebral blood flow, cerebral oxygenation, and brain function, emphasizing routine bedside transcranial Doppler ultrasound, cerebral oximetry, and continuous electroencephalography monitoring; shifting management strategies from mild hypothermia therapy to targeted temperature management, and defining the "446" target management pathway for the supercritical stage; emphasizing the assessment of static and dynamic cerebrovascular autoregulation functions through multimodal methods to achieve individualized optimal mean arterial pressure management; elevating cerebrospinal fluid management goals to the level of "glymphatic system" function maintenance; implementing a multidisciplinary collaborative, whole-process management model focusing on patients' long-term neurological functional outcomes; de-escalation criteria include multidimensional indicators such as recovery of brain structure, restoration of cerebrovascular autoregulation, improvement in cerebrospinal fluid dynamics, and reduction in biomarker levels; and integrating cutting-edge technologies like artificial intelligence into post-critical care management and rehabilitation planning. This consensus systematically integrates the entire process of neurocritical care management, reflecting the modern connotation of goal-oriented, dynamic, and multimodal integration in neurocritical care medicine. It aims to adapt to new trends such as deepening understanding of pathophysiological mechanisms, the integration of medicine and engineering, and the empowerment of artificial intelligence, thereby further advancing the discipline of critical care medicine.
4.Analysis on the practical challenges and paths of “clown doctors” practice from the perspective of narrative medicine
Hui WANG ; Yutong PAN ; Liqun LU ; Can ZHANG
Chinese Medical Ethics 2026;39(2):215-222
Narrative medicine focuses on empathy, relevance, and emotion, precisely aligning with the elements of “clown doctor” such as compassion, interaction, and pain relief. From the perspective of narrative medicine, the practice of “clown doctors” not only focuses on the emotional changes of patients but also enhances their sense of belonging by recreating their experiences. The key element for the success of “clown doctors” lies in establishing a multi-dimensional trust relationship among medical workers, patients, colleagues, and society, while ensuring their practice adheres to medical ethics norms. “Clown doctors” should concentrate on dimensions such as concept dissemination, clinical application, social recognition, and ethical practice of narrative medicine. They should also constantly optimize narrative techniques, deepen the understanding of patients’ stories, and intervene in the medical process in a more delicate and comprehensive way, thereby fostering in-depth communication and understanding between doctors and patients.
5.Comparison of the efficacy and safety of postauricular and intratympanic dexamethasone injection in the treatment of sudden sensorineural hearing loss with all frequency loss
Huidi PAN ; Qingfu BAO ; Changyang WANG
China Pharmacy 2026;37(5):639-644
OBJECTIVE To compare and analyze the clinical efficacy and safety of postauricular versus intratympanic dexamethasone injection in the treatment of sudden sensorineural hearing loss (SSNHL) with all frequency loss. METHODS A retrospective study was conducted on 150 patients with SSNHL with all frequency loss who were treated at the Hospital of Integrated Traditional Chinese and Western Medicine Affiliated to Zhejiang University of Chinese Medicine between June 1, 2020, and May 31, 2025. According to different injection routes, the patients were divided into the intratympanic group (intratympanic injection, n =78) and the postauricular group (postauricular injection, n =72). The differences in hearing threshold indicators and auditory outcomes before and after treatment, tinnitus/vertigo indicators, coagulation function indicators before and after treatment, as well as adverse reactions during treatment were compared between the two groups. RESULTS After treatment, both groups demonstrated significant reductions in the mean speech-frequency threshold and the mean hearing threshold compared with baseline ( P <0.05), whereas no statistically significant differences were observed between the two groups ( P >0.05). Therapeutic outcomes varied significantly across different degrees of hearing loss in each group (intratympanic group: P =0.005; postauricular group: P <0.001). Moreover, a significant linear trend relationship was observed between the degree of hearing loss and treatment efficacy grades ( P <0.001), indicating that the more severe the hearing loss, the poorer the treatment outcome ( P <0.05). Notably, the intratympanic group demonstrated superior efficacy compared with the postauricular group in patients with severe and profound hearing loss ( P <0.05). Among patients presenting with vertigo prior to treatment, the postauricular group exhibited a significantly higher response rate than the intratympanic group ( P <0.05), while no statistically significant difference was observed between the two groups in patients with tinnitus ( P >0.05). Regarding coagulation parameters, fibrinogen (FIB) levels and activated partial thromboplastin time decreased or shortened significantly follo wing therapy ( P <0.05), whereas thrombin time (TT) and prothrombin time were significantly prolonged ( P <0.05). Moreover, the postauricular group showed lower FIB levels and longer TT values compared with the intratympanic group ( P <0.05). No significant differences were identified between the two groups in the incidence of gastrointestinal reactions, bleeding manifestations, hepatic or renal dysfunction, or allergic reactions ( P >0.05); however, vestibular adverse reactions occurred more frequently in the intratympanic group ( P <0.05). CONCLUSIONS Compared with intratympanic injection, postauricular dexamethasone injection combined with conventional treatment regimens achieved comparable efficacy in overall hearing improvement in SSNHL patients with all frequency loss. However, the postauricular injection shows potential advantages in vertigo relief, enhancing treatment response in patients with varying degrees of hearing loss, regulating of coagulation function, and declucing of vestibular adverse reactions, while the intratympanic injection may be more suitable for patients with severe hearing loss.
6.Zuogui Wan Improve Ovarian Inflammatory Microenvironment and Stemness of Ovarian Germline Stem Cells in Ovarian Aging via cGAS/STING Signaling Pathway
Yunling ZHENG ; Xinyi PAN ; Zuang LI ; Yixuan WANG ; Junyi AN ; Yuxin ZOU ; Mengting XIAO ; Zheng CHEN ; Ling ZHU
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(7):1-10
ObjectiveTo investigate the mechanism of Zuogui Wan (ZGW) in improving ovarian inflammatory microenvironment and stemness of ovarian germline stem cells (OSCs) for treating ovarian aging via the cyclic guanosine monophosphate/adenosine monophosphate synthase (cGAS)/stimulator of interferon genes (STING) signaling pathway. MethodsForty C57BL/6 female mice were randomly divided into a blank group, a model group, a low-dose ZGW group (2.7 g·kg-1), a high-dose ZGW group (5.4 g·kg-1), and an estradiol valerate group (0.15 mg·kg-1), with 8 mice in each group. Except the blank group, all other groups received a single intraperitoneal injection of cyclophosphamide at 120 mg·kg-1 to establish an ovarian aging mouse model. After successful modeling, each group was continuously administered for 4 weeks, once daily. The physiological status of the mice was observed, and the ovarian index was calculated. The estrus cycle of the mice was monitored. Hematoxylin-eosin (HE) staining was used to observe pathological changes in ovarian tissue. Enzyme-linked immunosorbent assay (ELISA) was used to detect serum sex hormone levels. Serum inflammatory factors interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), and mouse interleukin-6 (IL-6) levels were detected using kits. Western blot was used to detect the protein expression of ovarian cGAS, STING, p-STING, TANK-binding kinase 1 (TBK1), p-TBK1, interferon-induced transmembrane protein 3 (Fragilis), and Vasa homolog protein (MVH). Quantitative real-time polymerase chain reaction (Real-time PCR) was used to detect the mRNA expression of inflammatory factors in ovarian tissue. Immunofluorescence double labeling was performed to locate OSCs in ovarian tissues, and fluorescence intensities of OSCs markers MVH and octamer binding transcription factor 4 (Oct4) were calculated. ResultsCompared with the blank group, the model group showed reduced body weight, ovarian wet weight, and ovarian index (P<0.01) and a disordered estrus cycle (P<0.01). In addition, the levels of serum follicle-stimulating hormone (FSH), TNF-α, IL-6, and IL-1β were increased (P<0.01), while anti-Müllerian hormone (AMH) and estradiol (E2) levels were decreased (P<0.01). The protein expression of cGAS, p-STING/STING, and p-TBK1/TBK1 in ovarian tissue was increased (P<0.05, P<0.01), while that of OSCs stemness factors MVH and Fragilis was reduced (P<0.01). Immunofluorescence indicated a reduction in MVH and Oct4 expression in OSCs (P<0.01). The mRNA expression of inflammatory factors TNF-α, IL-6, and IL-1β in ovarian tissue was increased (P<0.05, P<0.01). Compared with the model group, the treatment groups exhibited improved body weight, ovarian wet weight, and ovarian index (P<0.05) and a reduced rate of estrus cycle disorder (P<0.05, P<0.01). The levels of serum FSH, TNF-α, IL-6, and IL-1β were decreased (P<0.05, P<0.01), while AMH and E2 levels were increased (P<0.01). The protein expression levels of cGAS, p-STING/STING, and p-TBK1/TBK1 in ovarian tissue were decreased (P<0.05), while the protein expression of MVH and Fragilis was increased (P<0.05), and the fluorescence intensities of MVH and Oct4 were increased (P<0.05, P<0.01). The mRNA expression of inflammatory factors in ovarian tissue was decreased (P<0.05). ConclusionZGW alleviate ovarian inflammatory response, regulate ovarian microenvironment homeostasis, and maintain stemness of OSCs in ovarian aging mice probably by modulating the cGAS-STING signaling pathway, thereby improving ovarian function and delaying ovarian aging.
7.Diabetic Kidney Disease and Gut-kidney Axis: A Review
Yingchao WANG ; Yexin CHEN ; Hua ZHANG ; Jiangteng LIU ; Zhichao RUAN ; Xingru PAN ; Weijun HUANG ; Jinxi ZHAO
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(7):310-320
With the rising incidence of diabetes, diabetic kidney disease (DKD) has become a significant global health burden. Although current prevention and treatment strategies can partially delay the progression of DKD, the risk of patients advancing to end-stage renal disease remains high. Since the concept of the "gut-kidney axis" was first introduced at the International Congress on Dialysis in 2011, research on the role of gut microbiota in the pathogenesis of DKD has received increasing attention. This review summarizes the current research on gut microbiota, explores the mechanisms through which it contributes to DKD development, and outlines clinical approaches for DKD prevention and treatment based on the "gut-kidney axis" theory. Evidence indicates that dietary interventions, intake of probiotics or prebiotics, use of metformin and novel antidiabetic drugs, and application of traditional Chinese medicine (TCM) compound formulas can effectively improve gut microbiota composition, influence metabolite production, and restore the intestinal mucosal barrier. These interventions can further regulate intestinal innate immunity and inflammatory responses, thereby modulating the progression of DKD. Despite challenges posed by the traditional oral administration of water-decocted TCM compound formulas and the complexity of their ingredients, increasing evidence suggests that TCM may indirectly affect the occurrence and development of DKD by modulating gut microbiota. This finding provides a new perspective on the potential mechanisms of TCM in DKD treatment and may offer novel strategies for DKD prevention and therapy.
8.Identification and Analysis of bHLH Genes Related to Color Formation of Gastrodia elata Stem
Xue JIANG ; Dandan RAN ; Xiuwen WANG ; Xiaobo ZHANG ; Xiaohong OU ; Jie PAN ; Tao ZHOU ; Zhen OUYANG ; Jiao XU
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(8):202-209
ObjectiveGastrodia elata has evolved ecological types with shortened rhizome internodes and diversified flower and fruit coloration in response to different altitudes. Studying the genetic mechanisms of different ecotype germplasm is significant for guiding variety breeding in different cultivation areas. MethodsThe bHLH gene family was identified based on the whole-genome datasets of G. elata f. elata and G. elata f. glauca. Subsequently, the gene family members were subject to analysis, including gene structure, chromosomal localization, cis-acting elements, gene synteny, and phylogeny. Combined with transcriptome data and quantitative Real-time PCR, the expression patterns of bHLH genes in the stems of the different G. elata ecotype germplasm were analyzed. Finally, correlation analysis was conducted between gene expression patterns and color to obtain the key bHLH genes regulating the color formation of stem. ResultsA total of 63 bHLH genes were identified in both G elata f. elata and G. elata f. glauca, unevenly distributed across 17 chromosomes and clustered into 16 subfamilies, with significant expansion in some family members. Obvious inversions of bHLH genes on the same chromosome and interchromosomal translocations were detected in the two ecotype germplasm. Among these genes, 12 bHLH genes (such as bHLH62-3 and bHLH74) were associated with the bright yellow color of G elata f. elata stem, while 9 bHLH genes (such as PIL13, UNE12, and bHLH130) were correlated with the red color of G. elata f. glauca stem. Compared to G. elata f. glauca, the bHLH48 expression level was significantly higher in flowers and scale leaves of G elata f. elata, and the bHLH62-3 expression level was significantly higher in all organs of G elata f. elata. ConclusionsFunctional pathway divergence of the bHLH family members has occurred across different chromosomes in G elata f. elata and G. elata f. glauca. Through synergism or antagonism with other genes, 21 bHLH genes participate in the coloration metabolic pathway regulation of stems, flowers, and fruits. Specifically, bHLH62-3 is involved in regulating stem color differentiation in the anthocyanin biosynthesis pathway of G. elata, thus relevant to the color formation of stem. Additionally, GebHLH48 positively regulates flowering-related pathways to promote the early-flowering phenotype of G. elata f. elata. These findings have laid the foundation for analyzing the genetic regulatory mechanisms underlying the color formation of the G. elata stem.
9.Uncariae Ramulus Cum Uncis and Its Active Components in Treatment of Tourette Syndrome: A Review
Jiayu WANG ; Pan ZHENG ; Shaoyu LI ; Feng CHEN
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(8):337-346
Tourette syndrome (TS) is a highly prevalent neurodevelopmental disorder in children, clinically characterized primarily by motor and/or vocal tics. Its pathogenesis is associated with hyperactivity of the dopaminergic system in the basal ganglia, and current medical treatments are limited by adverse reactions and unsatisfactory efficacy. In traditional Chinese medicine (TCM), TS is classified under categories such as "liver wind" and "convulsions", and is considered to be closely related to liver dysregulation. Uncariae Ramulus Cum Uncis (URCU) is a commonly used wind-dispelling herb. URCU has a clearly defined origin and a rich chemical composition, with alkaloids as its major active constituents, including rhynchophylline and isorhynchophylline. Its plasma components include multiple prototype alkaloids, which exhibit metabolic differences and phenomena such as enterohepatic circulation. Its brain-entering components possess blood-brain barrier permeability, and their distribution is associated with pharmacological effects. In recent years, increasing numbers of studies have focused on the pharmacological effects and mechanisms of the active components of URCU in the treatment of TS. This article systematically reviews the mechanisms by which URCU and its main active constituents exert therapeutic effects on TS from the following aspects: regulation of monoamine and amino acid neurotransmitters to improve neurotransmitter system imbalance, neuroprotection and intervention in neuroinflammation-related pathways; antioxidant effects through activation of antioxidant signaling pathways, and immunomodulatory functions influencing immune cells and the gut microbiota. In addition, the clinical application of compound formulas containing URCU in the treatment of TS is summarized, with the aim of providing new perspectives for further research on the pharmacological mechanisms of URCU and the treatment of TS.
10.Identification and Analysis of bHLH Genes Related to Color Formation of Gastrodia elata Stem
Xue JIANG ; Dandan RAN ; Xiuwen WANG ; Xiaobo ZHANG ; Xiaohong OU ; Jie PAN ; Tao ZHOU ; Zhen OUYANG ; Jiao XU
Chinese Journal of Experimental Traditional Medical Formulae 2026;32(8):202-209
ObjectiveGastrodia elata has evolved ecological types with shortened rhizome internodes and diversified flower and fruit coloration in response to different altitudes. Studying the genetic mechanisms of different ecotype germplasm is significant for guiding variety breeding in different cultivation areas. MethodsThe bHLH gene family was identified based on the whole-genome datasets of G. elata f. elata and G. elata f. glauca. Subsequently, the gene family members were subject to analysis, including gene structure, chromosomal localization, cis-acting elements, gene synteny, and phylogeny. Combined with transcriptome data and quantitative Real-time PCR, the expression patterns of bHLH genes in the stems of the different G. elata ecotype germplasm were analyzed. Finally, correlation analysis was conducted between gene expression patterns and color to obtain the key bHLH genes regulating the color formation of stem. ResultsA total of 63 bHLH genes were identified in both G elata f. elata and G. elata f. glauca, unevenly distributed across 17 chromosomes and clustered into 16 subfamilies, with significant expansion in some family members. Obvious inversions of bHLH genes on the same chromosome and interchromosomal translocations were detected in the two ecotype germplasm. Among these genes, 12 bHLH genes (such as bHLH62-3 and bHLH74) were associated with the bright yellow color of G elata f. elata stem, while 9 bHLH genes (such as PIL13, UNE12, and bHLH130) were correlated with the red color of G. elata f. glauca stem. Compared to G. elata f. glauca, the bHLH48 expression level was significantly higher in flowers and scale leaves of G elata f. elata, and the bHLH62-3 expression level was significantly higher in all organs of G elata f. elata. ConclusionsFunctional pathway divergence of the bHLH family members has occurred across different chromosomes in G elata f. elata and G. elata f. glauca. Through synergism or antagonism with other genes, 21 bHLH genes participate in the coloration metabolic pathway regulation of stems, flowers, and fruits. Specifically, bHLH62-3 is involved in regulating stem color differentiation in the anthocyanin biosynthesis pathway of G. elata, thus relevant to the color formation of stem. Additionally, GebHLH48 positively regulates flowering-related pathways to promote the early-flowering phenotype of G. elata f. elata. These findings have laid the foundation for analyzing the genetic regulatory mechanisms underlying the color formation of the G. elata stem.

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