Effect of Lycium barbarum polysaccharide on functional recovery after decompression in a rat model of degenerative cervical myelopathy
Article information
Abstract
Study Design
Animal study.
Purpose
To investigate the effects of Lycium barbarum polysaccharide (LBP) on functional recovery following decompressive surgery in a rat model of cervical spondylotic myelopathy (CSM).
Overview of Literature
Surgical decompression halts the progression of CSM, but may leave residual neurologic deficits. LBP, derived from wolfberry, has been shown to modulate macrophage polarization and exert neuroprotective effects in acute spinal cord injury. We hypothesized that LBP would enhance functional recovery after surgical decompression in a rat model of CSM.
Methods
Female Sprague–Dawley rats with induced chronic cervical spinal cord compression were randomly assigned to the following groups: (1) LBP alone; (2) decompression alone; (3) decompression+LBP; (4) no treatment; and (5) sham surgery. Decompression was performed 4 weeks after compression. Outcomes were assessed using neurobehavioral tests, electrophysiology, and histological/immunohistochemical analyses.
Results
Baseline spinal cord injury severity was comparable across groups, as confirmed by electrophysiological studies. At 8 weeks postsurgery, rats in the decompression+LBP group showed significantly greater recovery of hindlimb function compared with rats in the decompression alone group. Improvements in somatosensory evoked potential and motor evoked potential latency and amplitude were also more pronounced in the decompression+LBP group. Histological analyses demonstrated reduced myelin vacuolation and increased myelin density with LBP treatment.
Conclusions
This study provides the first evidence that LBP derived from Chinese herbal medicine enhances neurological and functional recovery when combined with decompression in a rat model of CSM. These findings support further clinical investigation of LBP as an adjunctive therapy in patients with CSM.
Introduction
Cervical spondylotic myelopathy (CSM) is an age-related degenerative condition causing spinal cord compression, leading to progressive white matter demyelination and neurologic decline. Clinical manifestations include hand numbness, impaired upper-limb dexterity, and gait spasticity, which may progress to paralysis and incontinence [1]. Globally, the prevalence of spinal cord injury (SCI) has increased over the past 30 years, ranging from 236 to 1,298 cases per million across countries [2,3]. Surgical decompression is currently the only effective treatment for halting disease progression; however, approximately 45% of patients fail to achieve full recovery and continue to experience residual symptoms or disability postoperatively [4]. These limitations highlight the need for adjunctive strategies to improve outcomes for patients with CSM undergoing decompressive surgery.
Lycium barbarum (Wolfberry), used in Traditional Chinese Medicine for over 2,000 years for eye health and antiageing, contains Lycium barbarum polysaccharide (LBP), the main bioactive compound responsible for its therapeutic effects [5,6]. Recent studies suggest that Wolfberry exerts neuroprotective effects in animal models and humans with neurodegenerative conditions. LBP has shown efficacy in retinal disorders such as retinitis pigmentosa [7] and diabetic retinopathy [8], as well as direct neuroprotection in Alzheimer’s disease [9], Parkinson’s disease [10], and stroke [11]. Proposed mechanisms include reducing oxidative stress and apoptosis in retinal disease; lowering glutamate levels, amyloid-β accumulation, and homocysteine in Alzheimer’s disease; and inhibiting erythrocyte hemolysis and vascular smooth muscle cell migration via the MAPK signaling in stroke [12].
There is also growing evidence that LBP promotes recovery after neuronal injury. Studies suggest that LBP protects against postischemic neuronal death through activation of NR2A and inhibition of NR2B signaling pathways [13]. In addition, LBP has been shown to exert beneficial effects in a rat model of SCI, where delayed administration of LBP (7 days postinjury) reduced secondary damage by promoting M2 macrophage upregulation [14]. M2 macrophages are anti-inflammatory cells characterized by high interleukin-10 and transforming growth factor-β expression. Their polarization is believed to underlie the axon growth-promoting effects after SCI and is considered a desirable response [15].
In CSM, chronic cervical spinal cord compression can cause ischemia-reperfusion injury during decompression surgery, leading to oxidative damage and postoperative neurobehavioral decline. We hypothesized that LBP administration could mitigate this injury and enhance recovery when combined with surgical decompression for CSM. Wolfberry, widely consumed in Chinese communities as a safe oral supplement with no reported severe toxicity, offers strong translational potential for improving long-term recovery in CSM patients. This study aimed to investigate the synergistic effects of LBP supplementation alongside surgical decompression in a rat model of CSM.
Materials and Methods
Ethics statement
This study adhered to institutional animal care guidelines and was approved by the Shenzhen Peking University-HKUST Medical Center Animal Use and Care Committee (protocol number: 2021-115)
Animals
Forty adult female Sprague–Dawley (SD) rats (250–300 g) were housed under controlled conditions (12-hour light/dark cycle, 25°C). After 1 week of acclimatization, the rats were randomly assigned to five groups: (1) Control group (no compression); (2) Model group (surgery for spinal cord compression only); (3) LBP group (LBP 20 mg/kg, orally, once daily, from weeks 2 to 8 postcompression); (4) Decom group (surgical decompression alone at 4 weeks postcompression); and 5) LBP+Decom group (LBP supplementation as in the LBP group plus surgical decompression at 4 weeks).
Establishment of a rat chronic compression model
A previously described rat model of chronic compressive cervical injury was used [16]. After anesthesia with 3% pentobarbital (2 mL/kg, intraperitoneal injection), the occipital and nuchal regions were shaved and disinfected with iodine solution. A midline incision was made to expose the C5–C6 lamina, followed by removal of the ligamentum flavum to access the epidural space. A water-absorbing polymer (1×2×1 mm, 3% agarose gel; Amresco LLC, Solon, OH, USA) dried for 8 hours in a vacuum oven was implanted into the spinal canal at the C5 level. The polymer can absorb liquid in the spinal canal to reach its maximum expansion (fourfold in volume) in 2 hours. Once saturated with liquid, it remains stable at the maximal volume for 6 months, thereby producing chronic spinal cord compression. The incision was closed in layers, and animals were allowed to recover on a heated bed. Intramuscular penicillin was administered postoperatively to prevent infection [17].
Study design and intervention
Dried mature LB fruit was sourced from Zhongning, Ningxia Hui Autonomous Region, China [18]. LBP was prepared by diluting the weighted extract in ultrapure water (weight/volume) to prepare a stock solution (4 mg/mL). For the experimental group, LBP (20 mg/kg) was fed daily through a nasogastric tube beginning 1 week after the initial surgery and continuing until sacrifice.
Surgical decompression was performed at 4 weeks after model establishment according to group allocation. The procedure was similar to the establishment of cervical cord compression. After anesthesia and incision, the laminae from C4 to C6 were exposed. A laminectomy from C4 to C5 was performed, and the expanded polymer sheet was carefully removed. Muscle and skin were sutured after hemostasis. Animals were allowed to recover on a heating bed before being housed individually [19].
Neurobehavioral assessments
Neurobehavioral function was assessed using the forelimb locomotor assessment (FLA) and the Basso-Beattie-Bresnahan (BBB) locomotor score. These scores evaluate different combinations of rat joint movements, hindlimb movements, steps, forelimb and hindlimb coordination, trunk position and stability, paw placement, and tail position. The scores are awarded by independent observers while rats walked freely across an open surface [20].
The horizontal ladder walking test was used to quantify locomotor function. The device consisted of two Plexiglas walls (100×25 cm) with 25 stainless steel rods inserted as rungs, randomly adjusted at each time point. Trials were recorded using a video camera (HDR-CX290; Sony Inc., Tokyo, Japan), and the percentage of rungs used and paw placement errors were analyzed [21,22].
All animals were assessed at baseline, at weeks 2, 4, 6, and 8 using the ladder walking test, and by FLA and BBB scoring at day 3 and weekly thereafter. Body weight was also recorded.
Electrophysiological evaluation
Somatosensory evoked potentials (SEPs) were recorded to assess spinal cord function using our established protocol [23]. To elicit cortical SEP, tibial nerve stimulation was applied with a 0.2 ms square wave (5.1 Hz) at twice the motor threshold, defined as the lowest current eliciting a gastrocnemius twitch. During the SEP test, the effectiveness of nerve stimulation was evaluated by visual inspection of the twitch in the appropriate innervated muscle group. Cortical SEPs were recorded from the skull at Cz–Fz, amplified 100,000× with two amplifiers (SCXI-1120; National Instruments Co., Austin, TX, USA), and band-pass filtered (2–2,000 Hz). Signals were acquired with a DAQcard-1200 (National Instruments Co.) at 12-bit resolution and a 5,000 Hz sampling rate. To ensure SEP signal quality, a total of 500 SEP responses were averaged for each trial. Data were processed in MATLAB ver. 7.0 (Mathworks, Natick, MA, USA) on a Pentium 4 PC platform (3.2 GHz, 1 GB RAM). SEP latency and amplitude were analyzed.
Histological evaluations
At designated endpoints, rats were euthanized with intravenous sodium pentobarbital overdose. Perfusion was performed with 50 mL heparinized saline through the ascending aorta, followed by 300 mL formalin–picric fixative (4% formaldehyde, 0.4% picric acid in 0.16 mol/L phosphate buffer, pH 7.4). The entire cervical spinal cord was harvested, postfixed with 4% phosphate buffer liquid in formaldehyde solution for 72 hours, and embedded in paraffin. Transverse sections (5 μm) were stained with hematoxylin–eosin (H&E) and luxol fast blue (LFB). The slides were examined under a microscopic imaging system (FV-1000; Olympus, Tokyo, Japan).
Data analysis
Gait analysis, lesion morphometry, and immunohistochemistry were performed. Results are expressed as mean±standard deviation (SD). Group differences were assessed for statistical significance using one-way analysis of variance with Bonferroni post-hoc tests. Normality was verified using the Kolmogorov-Smirnov test before comparisons. A p-value <0.05 was considered indicative of statistical significance. SStatistical analysis were performed using IBM SPSS ver. 28.0 for Windows (IBM Corp., Armonk, NY, USA).
Results
Body weight
In the Control group, body weight increased steadily over 8 weeks. In contrast, all other groups experienced weight loss following chronic spinal cord compression. After decompression surgery, body weight began to recover; however, from weeks 5 to 8, the LBP group showed a significantly higher rate of weight gain than that in the Model and Decom groups (p<0.05). Within groups, compared to the lowest weights recorded 1-week postsurgery, the Model group showed significant recovery at 4 weeks (p<0.05), whereas LBP and LBP+Decom groups recovered significantly earlier, by week 3 (p<0.05) (Fig. 1).
Weight growth rate. Decom, decompression; LBP, Lycium barbarum polysaccharide; Preop, preoperative.
Behavioral tests
Fig. 2 shows the FLA and BBB scores for each group. Following chronic spinal cord compression, all four experimental groups exhibited a gradual decline. The Model group, which received no treatment postcompression, maintained poor FLA and BBB scores from week 4 onward. In the Decom group, decompression alone led to only modest improvements compared to the Model group. The LBP group showed mild recovery in FLA and BBB scores compared to the Model group, suggesting some neuroprotective effects of LBP; however, this alone was insufficient for substantial neurological improvement without decompression. The LBP+Decom group demonstrated the greatest functional gains, with significantly higher FLA and BBB scores among all groups, indicating marked functional improvement relative to either intervention alone.
The forelimb locomotor assessment (FLA) score (A) and Basso-Beattie-Bresnahan (BBB) (B) score. Decom, decompression; LBP, Lycium barbarum polysaccharide; Preop, preoperative.
Horizontal ladder
Usage rate of rungs: The average rung spacing was 4 cm. Because touches occurred more often with the forelimbs as opposed to the hindlimbs, there was a higher percentage of rungs used with the forelimbs (43.35%±7.23%) versus the hindlimbs (31.57%±3.94%).
Error rate of placements: Before surgery, rats crossed the ladder with minimal mistakes (forelimbs 2.25%±3.96%; hindlimbs 11.65%±8.63%).
After compression, both usage and error rates in all groups increased significantly at week 2, followed by gradual improvement over time. Compared with the Model and Decom groups, the LBP and LBP+Decom groups demonstrated lower usage and error rates (Table 1, Fig. 3).
Forelimb and hindlimb usage rate (%)
(A, B) Ladder test error rate. Decom, decompression; LBP, Lycium barbarum polysaccharide; Preop, preoperative.
Electrophysiological evaluation
The results of SEP and MEP evaluations are shown in Fig. 4 and Table 2. Following induction of slow chronic compression, the latency was significantly delayed and amplitudes were significantly decreased compared with baseline (p<0.01). Over time, both parameters showed a significant and distinct trend toward recovery in all four groups. Decompression and LBP administration each significantly reduced latency and increased amplitude (p<0.05), with comparable effects. The combination of surgery and LBP administration produced the greatest improvements, indicating a synergistic effect. These findings suggest that both interventions contributed to the restoration of spinal cord function, with the best recovery seen in the LBP+Decom group.
Somatosensory-evoked potentials (SEP) (A, B) and motor-evoked potentials (MEP) (C, D). Decom, decompression; LBP, Lycium barbarum polysaccharide; Postop, postoperative.
Statistical significance of changes in latency and amplitude SEP and MEP
Histological evaluations
In the Control group, the posterior funiculus (PF) and ventral horn (VH) displayed normal morphology with intact neuronal architecture (Fig. 5A, a–c). In the Model group, chronic compression caused significant neuronal loss in the VH (p<0.05) and vacuolation in the PF (p<0.05). Pathological features included degenerated white matter with vacuolated myelin, atrophic neurons with shrunken cytoplasm, and nuclear damage (Fig. 5A, d–f). Compared with the Model group, vacuolation and neuronal degeneration were reduced in the Decom group (Fig.5A, g–i) and more prominently in the LBP (Fig. 5A, j–l) and LBP+Decom groups (Fig. 5A, m–o). Both LBP and LBP+Decom groups showed significantly decreased vacuolation (p<0.05) and increased neuronal numbers (p<0.05), with restoration of abundant cytoplasm. The neuronal count was significantly higher in the LBP+Decom group than in the Decom group (p<0.05). No significant differences were found in neuronal count and vacuolation between the Decom and LBP groups or between the LBP and LBP+Decom groups.
(A–C) Lycium barbarum polysaccharide (LBP) reduce histopathology vacuolation and neuronal loss. (a, b, c) Normal white matter in posterior funiculus (PF) (a) and neuron morphology in ventral horn (VH) (c) of Control group; (d, e, f) histopathology vacuolation (d) in PF and neuronal loss in VH of Model group; (g, h, i) reduced histopathology vacuolation in PF (g) and neuronal degeneration (i) in VH of Decom group; significant decline of histopathology vacuolation in PF, and increase of neuron number and restoration of cellular architecture in LBP group (j, k, l) and especially LBP+Decom group (m, n, o). Values are presented as mean±standard deviation. Each group consisted of 8 (n=8/group). Decom, decompression; LBP, L. barbarum polysaccharide. A p-value <0.05 is statistically significant (*compared with Model group and **compared with Decom group).
LFB staining showed well-organized myelin fibers with clear grey–white matter boundaries in the control group (Fig. 6A, a–c). In the Model group, compression caused severe white matter loss with myelin staining and vacuolation (p<0.001) (Fig. 6A, d–f). Myelin staining area recovered and vacuolation decreased significantly in the Decom group (Fig. 6A, g–i), with greater recovery observed in the LBP (Fig. 6A, j–l) and LBP+Decom groups (p<0.05) (Fig. 6A, m–o). Compared with the Decom group, both the LBP and LBP+Decom groups showed superior restoration of myelin integrity (p<0.05), with relatively reorganized neural fibers. Among the treated groups, the LBP+Decom group exhibited the most pronounced improvements (p<0.05).
(A–C) Lycium barbarum polysaccharide (LBP) reduce myelin loss and promote remyelination in white matter. (a, b, c) Normal myelin staining in posterior funiculus (PF) (a) and anterior funiculus (AF) (c) of control group; (d, e, f) obvious decreased myelin staining area and severe vacuolation changes in PF (d) and AF (f) of model group; significant recovery of myelin staining area and decrease of vacuolation area were observed in Decom group (g, h, i), and especially in LBP group (j, k, l) and LBP+Decom group (m, n, o); (B, C) quantitative results of myelin staining area (B) and vacuolation area (C). Values are presented as mean±standard deviation. Each group consisted of 8 (n=8/group). Decom, decompression; LBP, L. barbarum polysaccharide. A p-value <0.05 is statistically significant (*compared with Model group, **compared with Decom group, and ***compared with LBP group).
Discussion
This study investigated the therapeutic effects of LBP in a rat model of chronic spinal cord compression, simulating CSM. Our results demonstrated that LBP therapy improved neurobehavioral function and electrophysiological parameters, while histological analyses revealed reduced axonal demyelination, fiber disruption, and neuronal necrosis. Notably, LBP combined with surgical decompression produced synergistic benefits, yielding superior motor function recovery, supported by both electrophysiological and histological evidence. Previous studies have shown that a subset of patients undergoing surgical decompression for CSM experience limited recovery or neurological deterioration. Retrospective analyses reported residual deficit or worsened function in 6.6%–11.6% of patients [24,25], while a prospective multicenter study in North America noted immediate postoperative neurological complications in 4% of patients and residual deficit or deterioration in neurological symptoms in 10% at 6-month follow-up [4]. One proposed mechanism is posterior cord shift after decompression, which may tether nerve roots [26]. In addition, surgical decompression for CSM can itself trigger ischemia-reperfusion injury with microglial recruitment and activation [27]. These factors highlight the clinical need for adjunctive therapies. Our findings suggest that LBP supplementation may act synergistically with decompression to enhance recovery and mitigate acute neurological deterioration in CSM.
This study has several limitations. First, mechanistic insights remain limited, as animals were sacrificed only at 8 weeks. Future studies should include histological analyses at earlier time points to examine microglia differentiation. Second, a dose-response relationship was not assessed as the selected dose was based on prior studies in rat models of neurodegenerative ophthalmic disease [28–30]. Third, rat neurobehavioral scoring systems such as FLA and BBB lack direct correlation with the minimal clinically important difference in human scales, making it uncertain whether the improvements observed here translate into clinically meaningful benefits. Nevertheless, given the lack of effective adjunctive strategies for enhancing surgical outcomes postdecompression in CSM, these findings offer promising translational potential.
Conclusions
In summary, LBP conferred neuroprotection in a rat model of CSM by reducing myelin loss and spinal cord cavitation. The combination of decompression surgery with LBP produced the greatest improvements in neurobehavioral and electrophysiological outcomes. Given its accessibility, favorable safety profile, and minimal side effects, Wolfberry-derived LBP represents a promising adjunctive therapy that warrants further evaluation in human clinical trials.
Key Points
Lycium barbarum polysaccharide (LBP) combined with surgical decompression produced synergistic benefits in a rat model of cervical spondylotic myelopathy (CSM).
LBP treatment improved neurobehavioral function and electrophysiological parameters.
Histological analyses revealed reduced axonal demyelination and neuronal necrosis with LBP treatment.
LBP represents a promising adjunctive therapy for CSM patients undergoing decompression surgery.
Notes
Conflict of Interest
No potential conflict of interest relevant to this article was reported.
Funding
This project is supported by Health Medical Research Fund (project no., 16172021), Hong Kong Government SAR.
Author Contributions
Conceptualization: KFS, YH, KYHK. Methodology: all authors. Data curation: KHW, GSL, RL, CYT. Formal analysis: KHW, GSL, RL, CYT. Investigation: KHW, GSL, RL. Project administration: KYHK. Resources: YH, KYHK. Funding acquisition: KYHK. Validation: YH, KYHK. Visualization: KFS, YH, KYHK. Supervision: YH, KYHK. Writing–original draft: KHW, GSL, RL, CYT. Writing–review & editing: KYHK. Final approval of the manuscript: all authors.