Astrocytes have been shown to produce several pro- and anti-inflammatory cytokines to maintain homeostasis of microenvironment in response to vast array of CNS insults. Some inflammation-related cytokines are responsible for regulating such cell events. Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine that can be inducibly expressed in the lesioned spinal cord. Unknown is whether MIF can facilitate the production of immunosuppressive factors from astrocytes to tune milieu following spinal cord injury. Following establishment of contusion SCI rat model, correlation of PGE2 synthesis-related protein levels with that of MIF was assayed by Western blot. ELISA assay was used to detect production of PGE2, TNF-α, IL-1β, and IL-6. Immunohistochemistry was performed to observe colocalization of COX2 with GFAP- and S100β-positive astrocytes. The primary astrocytes were treated by various inhibitors to validate relevant signal pathway. The protein levels of MIF and COX2, but not of COX1, synchronously increased following spinal cord injury. Treatment of MIF inhibitor 4-IPP to the lesion sites significantly reduced the expression of COX2, mPGES-1, and as a consequence, the production of PGE2. Astrocytes responded robustly to the MIF interference, by which regulated MAPK/COX2/PGE2 signal pathway through coupling with the CD74 membrane receptor. MIF-induced production of PGE2 from astrocytes was able to suppress production of TNF-α, but boosted production of IL-1β and IL-6 in LPS-activated macrophages. Collectively, these results reveal a novel function of MIF-mediated astrocytes, which fine-tune inflammatory microenvironment to maintain homeostasis. These suggest an alternative therapeutic strategy for CNS inflammation.

Astrocytes act as immune effector cells with the ability to produce a wide array of chemokines and cytokines in response to various stimuli. Macrophage migration inhibitory factor (MIF) is inducibly expressed in injured spinal cord contributing to excessive inflammation that affects motor functional recovery. Unknown is whether MIF can facilitate inflammatory responses through stimulating release of chemokines from astrocytes following spinal cord injury. Following the establishment of the contusion spinal cord injury rat model, the correlation of chemokine (C-C motif) ligand 5 (CCL5) expression with that of MIF was assayed by Western blot, ELISA, and immunohistochemistry. Immunoprecipitation was used to detect MIF interaction with membrane CD74 receptor. Intracellular signal transduction of MIF/CD74 axis was analyzed by transcriptome sequencing of primary astrocytes and further validated by treatment of various inhibitors. The effects of CCL5 released by astrocytes on macrophage migration were performed by transwell migration assay. The post-injury locomotor functions were assessed using the Basso, Beattie, and Bresnahan (BBB) locomotor scale. The protein levels of chemokine CCL5/RANTES were remarkably increased in the astrocytes of rat injured spinal cord, in parallel with the expression of MIF. Treatment of MIF inhibitor 4-IPP in the lesion sites resulted in a significant decrease of CCL5 protein levels. In vitro study revealed MIF was capable of facilitating CCL5 production of astrocytes through interaction with CD74 membrane receptor, and knockdown of this receptor attenuated such effects. Production of CCL5 in astrocytes was significantly blocked by inhibitor of c-Jun N-terminal kinase, rather than by those of ERK and P38. Recombinant CCL5 protein was found to be more effective in promoting migration of M2- compared to M1-type macrophages. Collectively, these data reveal a novel function of MIF in regulation of CCL5 release from astrocytes, which in turn favors for recruitment of inflammatory cells to the injured site of the spinal cord, in association with activation of excessive inflammation.

Background: Two activation states of reactive astrocytes termed A1 and A2 subtypes emerge at the lesion sites following spinal cord injury (SCI). A1 astrocytes are known to be neurotoxic that participate in neuropathogenesis, whereas A2 astrocytes have been assigned the neuroprotective activity. Heat shock transcription factor 1 (HSF1) plays roles in protecting cells from stress-induced apoptosis and in controlling inflammatory activation. It is unknown whether HSF1 is involved in suppressing the conversion of A1 astrocytes following SCI.Methods: A contusion model of the rat spinal cord was established, and the correlations between HSF1 expression and onset of A1 and A2 astrocytes were assayed by Western blot and immunohistochemistry. 17-AAG, the agonist of HSF1, was employed to treat the primary cultured astrocytes following a challenge by an A1-astrocyte-conditioned medium (ACM) containing 3 ng/ml of IL-1α, 30 ng/ml of TNF-α, and 400 ng/ml of C1q for induction of the A1 subtype. The effects of 17-AAG on the phenotype conversion of astrocytes, as well as underlying signal pathways, were examined by Western blot or immunohistochemistry.Results: The protein levels of HSF1 were significantly increased at 4 days and 7 days following rat SCI, showing colocalization with astrocytes. Meanwhile, C3-positive A1 astrocytes were observed to accumulate at lesion sites with a peak at 1 day and 4 days. Distinctively, the S100A10-positive A2 subtype reached its peak at 4 days and 7 days. Incubation of the primary astrocytes with ACM markedly induced the conversion of the A1 phenotype, whereas an addition of 17-AAG significantly suppressed such inducible effects without conversion of the A2 subtype. Activation of HSF1 remarkably inhibited the activities of MAPKs and NFκB, which was responsible for the regulation of C3 expression. Administration of 17-AAG at the lesion sites of rats was able to reduce the accumulation of A1 astrocytes.Conclusion: Collectively, these data reveal a novel mechanism of astrocyte phenotype conversion following SCI, and HSF1 plays key roles in suppressing excessive increase of neurotoxic A1 astrocytes.

Background: Reactive astrocytes are increasingly recognized as crucial regulators of innate immunity in degenerative or damaged central nervous system (CNS). Many proinflammatory mediators have been shown to drive inflammatory cascades of astrocytes through activation of NF-κB, thereby affecting the functional outcome of the insulted CNS. D-dopachrome tautomerase (D-DT), a newly described cytokine and a close homolog of proinflammatory macrophage migration inhibitory factor (MIF), has been revealed to share receptor and overlapping functional spectrum with MIF, but little is known about its roles in the neuropathological progression of the CNS and relevant regulatory mechanisms. Results: D-DT protein levels were significantly elevated within neurons and astrocytes following SCI. Analysis of transcriptome profile revealed that D-DT was able to activate multiple signal pathways of astrocytes, which converged to NF-κB, a hub regulator governing proinflammatory response. Rat D-DT recombinant protein was efficient in inducing the production of inflammatory cytokines from astrocytes through interaction with CD74 receptor. Activation of mitogen-activated protein kinases (MAPKs) and NF-κB was observed to be essential for the transduction of D-DT signaling. Administration of D-DT specific inhibitor at lesion sites of the cord resulted in significant attenuation of NF-κB activation and reduction of the inflammatory cytokines following SCI, and accordingly improved the recovery of locomotor functions. Conclusion: Collectively, D-DT is a novel proinflammatory mediator of astrocytes following SCI. Insights of its cell-specific expression and relevant proinflammatory mechanisms will provide clues for the control of CNS inflammation.

Background: With the advancement of surgical techniques and instruments, surgeries had been increasingly applied to patients with metastatic urothelial carcinoma. However, their survival benefits had not been carefully evaluated. Methods: Eligible articles were conducted by comprehensively searching three online databases (PubMed, EMBASE and Web of Science), published before May 1st, 2019. Overall survival (OS) and cancer-specific survival/progression-free survival (CSS/PFS) were analyzed to clarify their associations. Results: Finally, eight out of 3,581 articles were enrolled in this meta-analysis. In terms of OS, our results indicated that OS was positively associated with the patients underwent radical cystectomy (RC) (pooled HR =0.72, 95% CI, 0.64–0.81, P=0.158, I²=39.4%), but it was not significantly associated with the patients underwent metastasectomy (MC) (pooled HR =0.78, 95% CI, 0.56–1.08, P=0.093, I²=49.7%). As for CSS/PFS, our results displayed that patients could benefit from surgery (RC or MC) (pooled HR =0.56, 95% CI, 0.42–0.75, P=0.213, I²=35.3%). Conclusions: Despite the positive role of the RC in treating metastatic urothelial carcinoma, MC did not suggest a survival benefit in terms of OS. More strictly designed randomized controlled trials (RCTs) were needed to validate our findings.

Suprascapular nerve lesions caused by rotator cuff tears have been widely reported. However, the accuracy of electromyography as a definitive diagnostic method for nerve lesions is controversial. This study aims to evaluate the diagnostic effectiveness of electromyography for nerve lesions following varying degrees of rotator cuff tears in rat models. Twenty rats underwent bilateral rotator cuff tenotomy and were divided into four groups to simulate clinically relevant rotator cuff tears of different sizes. The supraspinatus tendon free ends in three groups were sutured at the tendon insertion site, the humeral head midpoint, and the coracoid process base. The fourth group underwent free retraction following supraspinatus and infraspinatus tendon transection. Nerve lesion incidence was assessed using in vivo electromyography under anesthesia and Neurofilament-Heavy immunofluorescence staining of the suprascapular nerve after sacrifice at 2 weeks postoperatively. The area under the receiver operating characteristic curve (AUC) for both single and combined electromyography indicators of nerve entrapment was < 0.7, indicating low diagnostic effectiveness. Immunofluorescence staining revealed a statistically significant positive correlation between nerve lesion incidence and tear size. Although widely used clinically, electromyography lacks sufficient diagnostic accuracy for suprascapular nerve lesions following rotator cuff tears. This underscores the need for surgeons to carefully evaluate suprascapular nerve integrity in such cases.