A highly biosimilar synthetic Calculus Bovis enhances cerebral blood flow and provides neuroprotection against ischemic stroke.

ETHNOPHARMACOLOGICAL RELEVANCE: Calculus Bovis (also known as ox bezoar, cattle gallstones, or Niu Huang in Chinese) has been used in traditional Chinese medicine (TCM) for centuries in China and Japan to treat a range of conditions, including high fever, convulsions, and stroke. However, the clinical use of natural Calculus Bovis (NCB) is limited due to its scarcity and high cost. As a result, several alternative substitutes have been developed in recent years, though their effectiveness remains unclear.

AIM OF THE STUDY: This study aimed to evaluate the efficacy of a highly biosimilar synthetic Calculus Bovis (HBSCB) in mitigating ischemic brain injury and enhancing cerebral blood flow (CBF) following ischemic stroke.

MATERIALS AND

METHODS

HBSCB was synthesized using synthetic biology, modeled after the formation mechanism of NCB. Chromatographic fingerprinting was conducted to compare the composition of HBSCB with that of NCB. Male C57BL/6J mice, subjected to transient middle cerebral artery occlusion (tMCAO), were administered varying doses of HBSCB (35.95, 71.9, and 143.8 mg/kg) via intragastric gavage, 1 h post-tMCAO. Therapeutic efficacy was assessed through measurements of brain infarction, neurological deficits and forelimb grip strength. Based on the results, 71.9 mg/kg was identified as the optimal dosage. This dose was then administered at 1, 4, and 6 h post-tMCAO to determine the therapeutic time window of HBSCB. Finally, Fluoro-Jade B (FJB) staining, Nissl staining, and laser speckle contrast imaging of CBF were utilized to investigate the neuroprotective mechanisms of HBSCB in mitigating ischemic brain injury.

RESULTS

HBSCB showed a high degree of similarity in key bioactive components with NCB, such as bilirubin and cholic acid. Administration of HBSCB at a dose of 71.9 mg/kg within 4 h post-tMCAO significantly reduced cerebral infarction and edema volumes, while also protecting neurons in the peri-infarct zone from cell death 3 days after tMCAO. Furthermore, HBSCB treatment notably alleviated neurological and motor deficits 1 day after tMCAO. These results demonstrate that HBSCB offers significant neuroprotective effects when administered within 4 h of tMCAO. Mechanistically, HBSCB significantly improved CBF in the ischemic region, which likely contributed to the restoration of energy levels and enhanced brain protection.

CONCLUSIONS

The present study demonstrates that HBSCB enhances CBF and provides neuroprotection against ischemic brain injury. These findings support the potential of HBSCB as a viable alternative to NCB for the treatment of ischemic stroke.