A 2025 bioRxiv preprint (Xue et al., doi:10.1101/2025.11.20.688774) proposed a causal cellular axis for the initiation of manual acupuncture: needle rotation winds collagen fibers at the acupoint, the transmitted tension stretches anchored fibroblasts, which release stem cell factor (SCF) and interleukin-33 (IL-33); SCF recruits mast cells and IL-33 activates them to secrete neuromodulatory molecules (substance P, serotonin), which excite sensory neurons — demonstrated only at the ST36 acupoint of mice with LPS-induced endotoxemia, where fibroblast depletion and IL-33 blockade abolish the anti-inflammatory and survival benefit of acupuncture and SCF/c-kit blockade suppresses mast-cell recruitment and degranulation at the acupoint. The question is whether this axis is in fact the general initiation mechanism of manual acupuncture — whether it operates at other acupoints, in other therapeutic contexts (pain, visceral disorders), and in humans — and how needle-evoked collagen tension is mechanistically coupled to SCF/IL-33 release (intracellular cascades, temporal dynamics, sensory-neuron subtypes engaged, and contributions of other immune cells). The SCF/IL-33 mediator chain of the axis has not been tested outside its original setting: fibroblast ablation at ST36 attenuates manual-acupuncture analgesia with reduced mast-cell degranulation in arthritic mice, but SCF and IL-33 were not examined there (Tu et al., Front Immunol 2026, doi:10.3389/fimmu.2026.1715313).

Background

Acupuncture is a component of traditional Chinese medicine in which fine needles are inserted at defined skin sites (acupoints) and manipulated, typically by rotation. ST36 (Zusanli), on the hindlimb below the knee, is among the most studied acupoints. Manual acupuncture (needle rotation without electrical current) must be distinguished from electroacupuncture, in which current passed through the needle drives nerves directly; the electroacupuncture mechanism is comparatively well characterized — for example, electroacupuncture at ST36 can drive an anti-inflammatory reflex from the vagus nerve to the adrenal glands with defined neuroanatomy (Liu et al., Nature 2021, doi:10.1038/s41586-021-04001-4) — whereas how the purely mechanical stimulus of traditional needle manipulation is converted into biological signals has remained unresolved.

Two research threads frame the problem. First, Helene Langevin and colleagues showed with ultrasound and microscopy that rotating an acupuncture needle pulls in and winds subcutaneous collagen fibers (Langevin et al., FASEB J 2002, doi:10.1096/fj.01-0925fje), and that acupuncture needling induces cytoskeletal remodeling in the fibroblasts anchored to that collagen matrix (Langevin et al., J Cell Physiol 2006, doi:10.1002/jcp.20623), establishing connective-tissue mechanotransduction — the conversion of mechanical force into biochemical signals — as a plausible initiation mechanism. Separately, mast cells (immune cells resident in skin and connective tissue, packed with granules of histamine, serotonin, and other mediators that they release upon degranulation) were repeatedly implicated in acupuncture's effects: mast cells are distributed along meridian-like lines in human skin at higher density near acupoints (Li 2019, doi:10.1142/S0192415X19500903), a preclinical meta-analysis found that skin mast cells contribute to acupuncture-induced analgesia in animals (Bae et al., J Pain 2021, doi:10.1016/j.jpain.2021.06.006), a mathematical model of mast-cell–nerve mechanotransduction at acupoints was published (Yao et al., Int J Biol Sci 2014, doi:10.7150/ijbs.8631), and peripheral mast cells were shown to be required for acupuncture's effects in a mouse model of Parkinson's disease (Oh et al., Front Aging Neurosci 2024, doi:10.3389/fnagi.2024.1376756). Whether and how the mechanical event and the mast-cell response were causally connected was, however, unknown.

Into this gap, Xue et al. (bioRxiv 2025, doi:10.1101/2025.11.20.688774) reported a complete causal chain in mice. During manual acupuncture at ST36, the needle winds collagen fibers (visualized by ultrasound, Masson's trichrome collagen stain, and collagen-I Western blot of needle-adherent material); selectively depleting fibroblasts at the acupoint abolishes acupuncture's protection in LPS-induced endotoxemia (lipopolysaccharide injection causing systemic inflammation, scored by serum IL-1β/IL-6/TNF-α and survival), its suppression of inflammation, and its activation of spinal neurons. In vitro, cyclic stretching of primary mouse skin fibroblasts induces release of SCF and IL-33 — pre-stored, rapidly secreted molecules — and this release is blocked by Y-27632, an inhibitor of ROCK (rho-associated coiled-coil kinase, a kinase that drives cellular actomyosin tension) (evidence from intracellular IL-33/SCF immunofluorescence intensity rather than a direct secretion assay), implicating cellular tension in the release mechanism (consistent with earlier work showing that mechanical strain induces IL-33 secretion from living murine fibroblasts in vitro and in vivo; Kakkar et al., J Biol Chem 2012, doi:10.1074/jbc.m111.298703). Of a panel of fibroblast-secreted factors, only SCF and IL-33 rose after stretch: SCF acts as a chemokine recruiting mast cells (blocked by ISCK03, an inhibitor of c-kit, the receptor tyrosine kinase for SCF on mast cells), while IL-33 activates mast cells to release substance P (SP) and serotonin (5-HT) (blocked by the soluble IL-33 receptor decoy sST2). Acupuncture-evoked signals reach the central nervous system, shown by c-fos expression (a standard marker of recent neuronal activation) in the lumbar dorsal horn of the spinal cord. Injecting IL-33 alone at the acupoint, without any needling, replicates acupuncture's protection against endotoxemia, and local blockade of IL-33 with sST2, like fibroblast depletion, eliminates that protection; the SCF/c-kit inhibitor ISCK03 was shown in vivo to suppress mast-cell accumulation and degranulation at the acupoint, while SCF injection alone did not reproduce the systemic benefit. The working model therefore couples needle mechanics to systemic therapeutic effect through a defined cellular axis.

The framework nonetheless rests on one experimental setting: one acupoint (ST36), one disease model (LPS endotoxemia in male C57BL/6J mice), in vitro substitution of a murine mastocytoma cell line (P815) for tissue mast cells, and pharmacological (rather than genetic) gain- and loss-of-function. The preprint itself states no limitations or open questions; the unresolved issues follow from its scope. Concurrent reviews by the same and independent groups (Sheng et al., Front Immunol 2026, doi:10.3389/fimmu.2026.1744045; Tu et al., Front Bioeng Biotechnol 2025, doi:10.3389/fbioe.2025.1662525) treat the mechanosensing framework as established at the hypothesis level but explicitly list the in vivo molecular details — such as how mechanically activated fibroblasts regulate immune-cell recruitment via IL-33-type signals — as open questions.

Problem Statement

Determine whether the fibroblast–SCF/IL-33–mast cell–sensory neuron signaling axis established at the ST36 acupoint in LPS-endotoxemic mice is the general initiation mechanism of manual acupuncture, and complete its mechanistic description. The overarching objective — testing and completing the proposed working model as the mechanistic framework of acupuncture initiation — comprises jointly linked questions: (1) Generality: does the same axis (collagen-mediated fibroblast stretch → SCF-dependent mast-cell recruitment and IL-33-dependent mast-cell activation → neuromodulator release → sensory-neuron activation → therapeutic effect) operate at acupoints other than ST36 and in therapeutic contexts other than acute systemic inflammation, such as pain and visceral disorders? Does it hold in human acupoint tissue, given species differences in skin thickness, collagen architecture, and immune-cell composition? (2) Mechanism: what intracellular signaling cascade (the ROCK-dependent tension pathway is implicated only by the Y-27632 observation) links fibroblast stretch to secretion of pre-stored SCF and IL-33; what are the temporal dynamics of this release and of mast-cell recruitment and degranulation at the acupoint; which sensory-neuron subtypes are engaged by the released substance P and serotonin (beyond bulk c-fos expression in the dorsal horn); and do other cell types, such as macrophages and dendritic cells, participate in the initiation cascade? Questions (1) and (2) are jointly posed and are to be settled on the existing evidence rather than on new laboratory data, so that any facet the evidence does not decide remains an open question. The contexts named ('such as pain and visceral disorders') delimit the class of therapeutic settings considered, and the human facet is bounded by the available human data and the limits of their comparability.

References

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