Zafirlukast in combination with pseudohypericin attenuates spinal cord injury and motor function in experimental mice
Authors: Fu Hua
Jin Sun
Yong-Yi Xu
Hai-Tao Yue
Xiao-Gang Chen
Shou-Guo Wang
Journal: Drug Design Development and Therapy (2018)
Discipline: PHARMACOLOGY & TOXICOLOGY
Keywords: 5-lipoxygenase; Cys-LT; mice; pseudohypericin; zafirlukast.
Affiliations: Department of Orthopedics, Huai'an First People's Hospital, Nanjing Medical University, Huai'an, Jiangsu 223300, People's Republic of China, ., Department of Gynaecology, Huai'an First People's Hospital, Nanjing Medical University, Huai'an, Jiangsu 223300, People's Republic of China.

Abstract:  Background: Biosynthesis of leukotriene (LT) by arachidonic acid involves 5-lipoxygenase (5-LO) as an important precursor. Here, we evaluated the role of pseudohypericin (PHP) for its postulated 5-LO inhibitory activity along with a Cys-LT receptor antagonist zafirlukast (ZFL) against inflammatory response and tissue injury in mice. Materials and methods: The spinal injury was induced by two-level laminectomy of T6 and T7 vertebrae. The inflammation was assessed by histology, inflammatory mediators by enzyme-linked immunosorbent assay, apoptosis by Annexin-V, FAS staining, terminal deoxynucleoti-dyltransferase-mediated UTP end labeling (TUNEL) assay and expression of Bax and Bcl-2 by Western blot. Effect on motor recovery of hind limbs was evaluated for 10 days postinjury. Results: The spinal injury resulted in tissue damage, apoptosis, edema, infiltration of neutrophils with increased expression of tumor necrosis factor-α (TNF-α) and cyclooxygenase-2 (COX-2). The spinal tissue showed elevated levels of prostaglandin E2 (PGE2), and LTB4 and increased phosphorylation of injured extracellular signal-regulated kinase-1/2 (ERK1/2). The PHP, ZFL and combination decreased inflammation, tissue injury and infiltration of neutrophils. Treatment also decreased the levels of PGE2, phosphorylation of extracellular signal-regulated kinase-1/2 (pERK 1/2), LT, TNF-α and COX-2 with a marked reduction in apoptosis and improved the motor function. Conclusion: The present study confirmed 5-LO antagonist activity of PHP and established its neuroprotective role along with ZFL. 

Retracted: No

PubPeer comments:
 Figure 3  Immunohistochemical localization and activity of MPO in spinal tissue of mice treated with PHP and ZFL their combination. Notes: In vehicle-treated control group mice, the tissue stained positive for MPO and presented increased MPO activity (A and F). Sham-operated group mice, tissue sections showed no positive staining for MPO and also had normalized MPO activity (B and F). MPO staining in tissues sections from PHP (C), ZFL (D) and their combination (E) a significant reduction in positive staining for MPO activity was seen compared to vehicle-treated control group. ![file] What happened in panel A? ![file] What happened in C and D? (contrast increased for clarity). Fundamentally they re the same (e.g. around the edges) but with repeated elements. Some of these have been vertically or horizontally flipped between the two panels, ![file] , 
 Fig 1.  Effect of pseudohypericin and zafirlukast on histological changes on the spinal cord tissue 24 h after inducing spinal injury. Notes: (A) Vehicle-treated control group showing occurrence of edema and changes in white matter of perilesional zone. (B) Sham-vehicle-treated group mice showing no signs of changes. The severity of spinal cord injury decreased in tissues with reduction in edema and preservation of white matter observed in sections (C) pseudohypericin- and (D) zafirlukast-treated mice. The results were improved in (E) after combining pseudohypericin and zafirlukast. ![file] (A). I have concerns. ![file] (C) and (E) have a lot in common. Here dramatised by color-reversing (E) and superimposing it on (C) with 50% transparency so that shared background cancels out. ![file] (C) in isolation: ![file] , 
 Fig 7.  TUNEL assay of spinal cord tissue of mice subjected to spinal injury followed by treatment of pseudohypericin, zafirlukast and their combination. Notes: The tissue harvested after 24 h of injury and received treatment of vehicle only showed appearance of brown color cells suggesting apoptosis (A). In contrast the tissues of mice treated with pseudohypericin (C), zafirlukast (D) and their combination (E) demonstrated reduction in number of apoptotic cells, no apoptotic fragments were detected in sham-operated vehicle-treated mice spinal tissues (B). TUNEL-terminal deoxynucleotidyltransferase-mediated UTP end labeling assay. The arrows indicate the TUNEL positive cells. ![file] (E) is fundamentally a horizontally-flipped version of (C) [purple boxes], though with the graphic elements shuffled around on the background. The other Figures also reward close scrutiny but I have lost the will to go on. , 
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