Pirfenidone inhibits fibroblast proliferation, migration or adhesion and reduces epidural fibrosis in rats via the PI3K/AKT signaling pathway.

OBJECTIVE: This present study aims to assess the effect of pirfenidone (PFD) on inhibiting fibroblast proliferation, migration or adhesion in vitro and reducing laminectomy-induced epidural fibrosis in vivo. METHODS: The effect of PFD on proliferation inhibition was evaluated with flow cytometry, CCK-8, EdU and western-blotting assays. Altered properties in migration and adhesion were confirmed by wound-scratch, transwell, immunofluorescence (IF), cell adhesion and western-blotting assays. Additionally, fifty male healthy Sprague-Dawley rats were subjected to laminectomy and then treated with various concentrations of PFD. After 4 weeks, the degree of epidural fibrosis was evaluated by histological analysis. RESULTS: In vitro, the results of flow cytometry, CCK-8, EdU and western-blotting assays showed that PFD reduced fibroblast proliferation by a dose-dependent manner. And the results of wound-scratch, transwell, IF, cell adhesion and western-blotting assays showed that the migration and adhesion of fibroblasts could be inhibited and the cytoskeleton could also be altered in a dose-dependent manner. And the inhibitory effect of PFD could be partially reversed in the PI3K overexpression experiment, which indicated that the capability of PFD to inhibit fibroblast proliferation, migration and adhesion might be through the PI3K/AKT signaling pathway. In vivo, an obvious reduction in epidural fibrosis was observed in groups topically treated with PFD. CONCLUSIONS: Topical PFD application obviously suppressed laminectomy-induced epidural fibrosis, possibly by inhibiting fibroblast proliferation, migration and adhesion via the PI3K/AKT signaling pathway. PFD may be a safe and effective pharmaceutical to reduce clinical epidural fibrosis.

MICAL2 regulates myofibroblasts differentiation in epidural fibrosis via SRF/MRTF-A signaling pathway.

AIM: To determine the role of MICAL2 in myofibroblasts differentiation and epidural fibrosis. BACKGROUND: Epidural fibrosis (EF) may develop following laminectomy and aberrant myofibroblasts differentiation and excessive extracellular matrix (ECM) accumulation play key roles in the formation of EF. Dense epidural fibrosis results to the poor surgical outcomes and failed back surgery syndrome (FBSS), and there is no effective treatment available. Molecule interacting with Casl2 (MICAL2) has been demonstrated to participate in multiple cellular processes by regulating actin cytoskeleton dynamics. However, its role in epidural fibrosis remains totally unverified. MATERIALS AND METHODS: The potential functions and mechanisms of MICAL2 were explored using western blotting, immunofluorescence and lentivirus infection. KEY FINDINGS: In our study, we determined that the MICAL2 expression was elevated in epidural fibrotic tissues and TGF-ß1-stimulated fibroblasts. Moreover, knockdown of MICAL2 using MICAL2-specific short hairpin RNA attenuated TGF-ß1-induced myofibroblasts differentiation and epidural fibrosis both in vitro and vivo, as indicated by decreased scar formation, reduced collagen production and down-regulated expression of α-SMA, collagen-1 and fibronectin. We also demonstrated that MICAL2 knockdown affected the migratory capability of fibroblasts in vitro. By further mechanistic research, we revealed that the MRTF-A nuclear translocation was inhibited in response to the knockdown of MICAL2 in fibroblasts and MICAL2 served as a pro-fibrotic factor in an SRF/MRTF-A-dependent manner. SIGNIFICANCE: In conclusion, our results indicated that MICAL2 mediated myofibroblasts differentiation and promoted epidural fibrogenesis via SRF/MRTF-A signaling pathway, suggesting manipulation of MICAL2 activity as a novel alternative strategy for the prevention of epidural fibrosis.

The peridural membrane of the spine has characteristics of synovium.

The peridural membrane (PDM) is a well-defined structure between dura mater and the wall of the spinal canal. The spine may be viewed as a multi-segmented joint, with the epidural cavity and neural foramina as joint spaces and PDM as synovial lining. The objective of this investigation was to determine if PDM has histological characteristics of synovium. Samples of the PDM of the thoraco-lumbar spine were taken from 23 human cadavers and analyzed with conventional light microscopy and confocal microscopy. Results were compared to reports on similar analyses of synovium in the literature. Histological distribution of areolar, fibrous, and adipose connective tissue in PDM was similar to synovium. The PDM has an intima and sub-intima. No basement membrane was identified. CD68, a marker for macrophage-like-synoviocytes, and CD55, a marker for fibroblast-like synoviocytes, were seen in the lining and sub-lining of the PDM. Multifunctional hyaluronan receptor CD44 and hyaluronic acid synthetase 2 marker HAS2 were abundantly present throughout the membrane. Marked presence of CD44, CD55, and HAS2 in the well-developed tunica muscularis of blood vessels and in the body of the PDM suggests a role in the maintenance and lubrication of the epidural cavity and neural foramina. Presence of CD68, CD55, and CD44 suggests a scavenging function and a role in the inflammatory response to noxious stimuli. Thus, the human PDM has histological and immunohistochemical characteristics of synovium. This suggests that the PDM may be important for the homeostasis of the flexible spine and the neural structures it contains.

Effective delivery of mitomycin-C and meloxicam by double-layer electrospun membranes for the prevention of epidural adhesions.

Epidural adhesion between the spinal dura and the surrounding fibrous tissue often occurs post-laminectomy, resulting in clinical symptoms such as nerve compression and severe pain. In this study, we report a drug-loaded double-layered electrospun nanofiber membrane to prevent the occurrence of epidural adhesion. The nanofibers in both layers are made of a mixture of polycaprolactone (PCL) and chitosan (CS) but at different weight ratios. The bottom layer contacting to the spinal dura is loaded with meloxicam (MX) to prevent inflammation. The top layer that contacts to the fibrous tissue is doped with mitomycin-C (MMC) to inhibit the synthesis of DNA and collagen. The two types of drugs are released from the double-layered membrane within about 12 days. Meanwhile, the membrane can inhibit fibroblasts proliferation in vitro while show no cytotoxicity. In a rabbit laminectomy model, the double-layered membrane can effectively prevent the epidural adhesion formation based on the adhesion scores, histological and biochemical evaluations. The combination release of MX and MMC can signally reduce the inflammation reaction and collagen I/III expression relative to the case with the membranes loaded with only either one type of the drugs. This approach offers new progresses in constructing dual drug delivery system and provides innovative barrier strategy in inhibiting epidural adhesion post-laminectomy.

Emodin promotes fibroblast apoptosis and prevents epidural fibrosis through PERK pathway in rats.

BACKGROUND: Laminectomy is usually classed as a common orthopedic surgery, but postoperative epidural fibrosis often leads to less-than-desirable clinical outcomes. As demonstrated by prior studies, emodin (EMO) exerts an anti-fibrotic effect. Here, we carried out investigation into the inhibitory effect created by EMO application on epidural fibrosis after laminectomy in rats. METHODS: The paper conducts a series of experiment. In vitro, we observed the effect of EMO on fibroblasts by Cell Counting Kit-8 (CCK-8) assay. Apoptosis of fibroblasts induced by EMO was detected by western blot, TUNEL assay, and flow cytometry. The results revealed that EMO was capable of inducing fibroblast apoptosis, and the proteins of PERK pathway also changed accordingly. In vivo, the effect of EMO on epidural fibrosis in 12 male Sprague-Dawley rats was observed by histological staining. RESULTS: CCK-8 assay indicated that EMO was effective in reducing fibroblast viability in a time- and a dose-dependent manner. TUNEL assay and flow cytometry analysis have demonstrated that the apoptotic rate of fibroblasts increased as the EMO concentration rose. Western blot analysis proved that EMO promoted the relative expression of p-perk and p-eIF2α and that the expression of its downstream proteins CHOP and GRP78 was also enhanced. The expression of apoptotic protein Bax and cleaved PARP was upregulated, whereas the expression of anti-apoptotic protein Bcl-2 was downregulated. In addition, histological and immunohistochemical analysis demonstrated that EMO functioned to inhibit epidural fibrosis and increase GRP78 expression in fibrous tissue by promoting apoptosis of fibroblasts. CONCLUSIONS: EMO could have inhibitory effect on epidural fibrosis in a concentration-dependent manner. The potential mechanism might be through PERK signaling pathway to promote fibroblast apoptosis. It has a possibility to be taken as a novel method for the treatment of epidural fibrosis.

Apigenin inhibits fibroblast proliferation and reduces epidural fibrosis by regulating Wnt3a/ß-catenin signaling pathway.

BACKGROUND: Failed back surgery syndrome (FBSS) is a common complication after the laminectomy. Epidural fibrosis is the major cause of lower back pain and other complications. Numerous studies have shown that apigenin (API) could treat various fibrotic diseases by regulating various signaling pathways, whereas no study has discussed whether API can inhibit fibroblast proliferation and reduce epidural fibrosis after the laminectomy by regulating Wnt3a/ß-catenin signaling pathway. METHODS: Human fibroblasts were cultured and treated with API in different concentrations for 24 h. CCK-8 detection and EdU incorporation assay were performed to detect cell viability and cell proliferation. Western blotting analysis was applied to detect expressions of proliferative proteins, Wnt3a, and its downstream proteins. Moreover, the Wnt3a gene was overexpressed in fibroblasts to define the relationship between Wnt3a/ß-catenin signaling pathway and fibroblast proliferation. Wnt3a overexpressed fibroblasts were treated with API to verify if it could reverse the effects of API treatment. Twenty-four Sprague-Dawley rats were randomly divided into four groups. Laminectomy was performed and the rats were gavaged with different doses of API or 5% sodium carboxyl methyl cellulose (CMC-Na) solution for 1 month. The abilities of API to inhibit fibroblast proliferation and to reduce epidural fibrosis were evaluated using histological and immunohistochemical analysis. RESULTS: CCK-8 detection and EdU incorporation assay demonstrated that API could inhibit the viability and proliferation rate of fibroblasts in a concentration-dependent manner. The Western blotting analysis revealed that API could inhibit the expressions of PCNA, cyclinD1, Wnt3a, and its downstream proteins. The overexpression of Wnt3a in fibroblasts could upregulate the expressions of proliferative proteins such as PCNA and cyclinD1. The inhibitory effect of API on PCNA, Wnt3a, and its downstream proteins was partially reversed by overexpression of Wnt3a. Moreover, the results of the histological and immunohistochemical analysis revealed that API could reduce the epidural fibrosis in rats by inhibiting fibroblast proliferation in a dose-dependent manner. CONCLUSIONS: API can inhibit fibroblast proliferation and reduce epidural fibrosis by suppressing Wnt3a/ß-catenin signaling pathway, which can be adopted as a new option to prevent epidural fibrosis after the laminectomy.

Effect of Cetuximab on the Development of Epidural Fibrosis Based on CD105 and Osteopontin Immunohistochemical Staining.

STUDY DESIGN: The effect of cetuximab on the development of epidural fibrosis (EF) was assessed using immunohistochemical methods as well as antibodies for CD105 and osteopontin (OPN). OBJECTIVE: The goal of this study was to assess of EGFR inhibition for the postoperative treatment of fibrosis. SUMMARY OF BACKGROUND DATA: EF is one of most common causes of failed back surgery syndrome, which occurs after laminectomy. Numerous causes and mechanisms have been proposed to explain its development after laminectomy. Many agents have been tested to prevent the development of EF. EGFR, a multi-functional transmembrane glycoprotein, causes cell growth, proliferation, and EF by interacting with epidermal growth factor and TGF-ß1. The inhibition of postoperative fibrosis using cetuximab, an epidermal growth factor receptor blocker, is theoretically possible. However, this has not been tested to date. METHODS: Sixteen Wistar-Albino rats were divided into two groups, namely, control and cetuximab groups. L1-2 laminectomy alone was performed in both groups, and topical cetuximab was applied to the treatment group. After 6 weeks, rats were sacrificed and examined histopathologically and immunohistochemically; EF tissue was also graded. Statistical significance was accepted at P < 0.05. RESULTS: Fibroblast counts and fibrosis density, determined by histopathologic examination, and EF, according to immunohistochemical assessment based on CD105, were found to be higher in the treatment group than in the control group, and this was statistically significant (P < 0.001). Based on OPN staining, the results were consistent with classical methods, and no significant difference was detected among the groups (P = 0.358). CONCLUSION: Our study revealed that cetuximab inhibits the development of EF and that CD105, and not OPN, is a reliable marker for grading EF. In addition, cetuximab did not result in toxic, systemic side effects in surrounding tissues. LEVEL OF EVIDENCE: N/A.

Mechanical barriers and transforming growth factor beta inhibitor on epidural fibrosis in a rabbit laminectomy model.

BACKGROUND: TGF-ß has been described as a mediator of fibrosis and scarring. Several studies achieved reduction in experimental scarring through the inhibition of TGF-ß. Fibroblasts have been defined as the cell population originating fibrosis, blocking fibroblast invasion may impair epidural fibrosis appearance. For this purpose, biocompatible materials used as mechanical barriers and a TGF-ß inhibitor peptide were evaluated in the reduction of epidural fibrosis. METHODS: A L6 laminectomy was performed in 40 New Zealand white rabbits. Divided into four groups, each rabbit was assigned to receive either collagen sponge scaffold (CS group), gelatin-based gel (GCP group), P144® (iTGFß group), or left untreated (control group). Four weeks after surgery, cell density, collagen content, and new bone formation of the scar area were determined by histomorphometry. Two experienced pathologists scored dura mater adhesion, scar density, and inflammatory infiltrate in a blinded manner. RESULTS: In all groups, laminectomy site was filled with fibrous tissue and the dura mater presented adhesions. Only GCP group presented a significant reduction in collagen content and scar density. CONCLUSION: GCP treatment reduces epidural fibrosis although did not prevent dura mater adhesion completely.

Homoharringtonine prevents surgery-induced epidural fibrosis through endoplasmic reticulum stress signaling pathway.

Surgery-induced epidural fibrosis after laminectomy often results in poor clinical outcomes. Fibroblasts proliferation is considered to be one of the major causes of epidural fibrosis formation. Previously, there was no research about the effect of Homoharringtonine(HHT) on inhibiting fibroblast proliferation and reducing epidural fibrosis. Here, we performed in vitro and in vivo experiments to explore the effect of HHT on inhibiting fibroblast proliferation, inducing fibroblast apoptosis and preventing epidural fibrosis formation. In vitro, the effect of HHT on inhibiting fibroblasts was detected by CCK-8 assay. Besides, the effect of HHT on causing fibroblast apoptosis was investigated via Western blots, flow cytometry and TUNEL assay. Results suggested that HHT could inhibit fibroblasts proliferation and induce apoptosis. And the marker proteins of endoplasmic reticulum (ER) stress were also changed positively. In vivo, histological macroscopic assessment, hydroxyproline content analysis and histological staining were used to detect the effect of HHT on reducing epidural fibrosis. The results showed that HHT had positive suppressive effects on epidural fibrosis following laminectomy in rats. TUNEL assay in epidural tissue suggested that HHT could obviously induce fibroblasts apoptosis. Immunohistochemistry staining showed that the expression of two important ER stress markers(78-kDa glucose-regulated protein and C/EBP homologous protein) were also increased. In conclusion, this research showed that HHT could reduce epidural fibrosis formation after laminectomy, and the potential mechanism might through inhibiting fibroblasts proliferation and inducing fibroblasts apoptosis via ER stress signaling pathway. It might provide a novel agent for reducing epidural fibrosis after laminectomy surgery.

Cervical Epidural Contrast Spread Patterns in Fluoroscopic Antero-Posterior, Lateral, and Contralateral Oblique View: A Three-Dimensional Analysis.

Objective: To describe and to analyze cervical epidural contrast patterns seen in antero-posterior (AP), contralateral oblique (CLO), and lateral view. To identify factors that might help in predicting contrast distribution pattern and extent. Method: Spread of contrast in the cervical epidural space was prospectively studied in AP, lateral, and three CLO views. Results: CLO view showed contrast spread of variable thickness with its posterior margin overlying the ventral interlaminar line (VILL). In the lateral view, the spread was also of variable thickness, but the posterior margin of the contrast lay on the spinolaminar line in only 10 of 24 patients. Ventral contrast spread was not visualized in any patient. In the AP view, bilateral spread was seen in 14 of 24 subjects, and nerve root spread was seen in 16 of 24 subjects. No association of the pattern of spread or dispersion was seen to patient age, volume injected, or needle location. Conclusions: The CLO view provides a consistent radiological landmark for the posterior margin of contrast in the dorsal epidural space; the lateral view fails to provide such a consistent landmark. The thickness of the spread is variable, both in the CLO and in the lateral view. Thick spread extending into the foramen in the CLO view and over the articular pillars in the lateral view is frequent and should not be misconstrued as subdural or intrathecal spread. In contradistinction to previous studies, true ventral spread was not seen in any patient. When using low volumes, contrast spread is independent of patient age, volume injected, or needle tip location in the AP view.

Hemostasis vs. epidural fibrosis?: A comparative study on an experimental rat model of laminectomy.

AIM: The aim of this study was to evaluate the histopathological and biochemical impact and effectiveness of two hemostatic agents, Ankaferd blood stopper (ABS) and Microporous Polysaccharide Hemospheres (MPH), on epidural fibrosis in an experimental rat laminectomy model. MATERIAL AND METHODS: Twenty adult Wistar albino rats were divided into MPH-treated (n=6), ABS-treated (n=6) and control (n=8) groups. Laminectomy of the lumbar spine was performed in all animals and treatment groups were exposed to MPH and ABS while closure was applied in control group as per usual. Epidural fibrosis was evaluated in all groups macroscopically, histopathologically, biochemically and with electron microscopy four weeks later. RESULTS: Statistically, it was found that MPH-treated group had significantly less epidural fibrosis compared to ABS-treated and control groups. CONCLUSION: We compared two hemostatic agents for their propensity to cause adhesions in the present study. Our results show that MPH significantly reduces epidural scar formation and dural adhesion in a rat model of laminectomy while ABS increases postoperative fibrosis.

Anti-adhesive effect of poloxamer-based thermo-sensitive sol-gel in rabbit laminectomy model.

Poloxamer-based thermo-sensitive sol-gel has been developed to reduce the incidence of postoperative scar formation at the laminectomy site. The purpose of this study was to evaluate the anti-adhesive effect of poloxamer based thermo-sensitive sol-gel compared to hyaluronate based solution after laminectomy, using a rabbit model. A thermo-sensitive anti-adhesive with a property of sol-gel transition was manufactured by a physical mixture of Poloxamer188/407, Chitosan and Gelatin. The viscosity in different temperatures was assessed. 72 adult New Zealand rabbits underwent lumbar laminectomy and were randomly divided into experimental (treated with the newly developed agent), positive (treated with hyaluronate based solution), and negative control groups. Each group was subdivided into 1 and 4-week subgroups. Gross and histological evaluations were performed to assess the extent of epidural adhesion. The experimental group showed significantly higher viscosity compared to the positive control group and showed a significant increase of viscosity as the temperature increased. Gross evaluation showed no statistically significant differences between the 1- and 4-week subgroups. However, histologic evaluation showed significant differences both in 1- and 4-week subgroups. Although the 4-week histologic results of the experimental and the positive control subgroups showed no significant difference, both subgroups revealed higher value compared to the negative control subgroup with regard to the ratio of adhesion less than 50 %. The new poloxamer based thermo-sensitive agent showed superior efficacy over the hyaluronate based agent at 1 week postoperatively. At 4 weeks postoperatively, there were no statistically significant differences between the two agents, although both showed efficacy over the sham group.

Comparison of Intrathecal Concentrations of Acyclovir following Epidural and Intravenous Administration in Rats.

BACKGROUND: Herpes zoster is a disease caused by reactivation of varicella-zoster virus in sensory cranial nerves and dorsal root ganglion. Our presumption was that epidural administration of acyclovir near the viral burden could be more advantageous than intravenous (IV) administration. The cerebrospinal fluid concentration of acyclovir after epidural administration was determined to be higher than that after IV administration in rats. OBJECTIVE: In this study, we tested the hypothesis that the concentration of acyclovir in CSF after epidural administration is higher than that achieved after IV administration in rats. STUDY DESIGN: A randomized controlled animal trial. METHODS: A total of 30 adult male Sprague-Dawley rats were used. The rats were randomly divided into 2 equal groups, epidural (Group Epi) and IV (Group IV) administration groups (n = 15). Group Epi was further subdivided into 3 groups according to acyclovir dosage; each group comprised 5 animals receiving injections at dosages of 0.3 mg, 0.6 mg, and 0.9 mg. Group IV was also subdivided into 3 groups receiving dosages of 3 mg, 6 mg, and 9 mg. We measured CSF and plasma acyclovir concentrations one hour after administration. RESULTS: In Group Epi, the median plasma concentrations of acyclovir were lower than that in CSF (P < 0.05). In Group IV, the median plasma concentrations of acyclovir were significantly higher than that in CSF (P < 0.05). The CSF concentrations of acyclovir in Group Epi were significantly higher than that in Group IV (P < 0.05). The plasma concentrations of acyclovir in Group Epi were significantly lower than that in Group IV (P < 0.05). LIMITATIONS: There were no references of equivalent dosages of acyclovir between IV and epidural administration. However, it is obvious in this study that epidural administration of a low dose of acyclovir can more effectively increase its concentration in the intrathecal space than IV administration. CONCLUSIONS: Epidural administration of acyclovir provides superior drug concentrations in the intrathecal space compared to IV administration. KEY WORDS: Acyclovir, epidural injection, herpes zoster, varicella zoster virus.

Taurine Reduced Epidural Fibrosis in Rat Models after Laminectomy via Downregulating EGR1.

BACKGROUND/AIMS: Epidural fibrosis, a common complication after laminectomy, has been demonstrated to be closely associated with poor surgical outcomes. Previous studies showed that taurine had remarkable anti-fibrotic effects on lung and liver fibrosis. We performed this study to investigate the effects of taurine in rat models of epidural fibrosis after laminectomy and to explore the potential molecular mechanism. METHODS: Laminectomy was performed on each rat to establish epidural fibrosis model. After taurine treatment, Masson's trichrome and immunohistochemistry staining were used to examine epidural fibrosis. Cell viability was determined using the Cell Counting Kit-8 assay. Annexin V/Propidium Iodide double staining was performed to detect fibroblasts apoptosis. Microarray was adopted to identify significantly changed mRNAs. mRNA expression was measured by qRT-PCR. Lentivirus infection was performed to establish stable knockdown and overexpression cell lines. The expression of fibrosis-related proteins was determined via Western blot. RESULTS: Taurine treatment markedly reduced laminectomy-induced epidural fibrosis in rat models. However, this effect of taurine was independent on TGF-ß/Smad pathway, evidenced by no change in the expression of TGF-ß and its receptors. Besides, taurine had almost no effect on cell apoptosis. Interestingly, taurine treatment significantly decreased expression of EGR1 (Early growth response protein 1), an enhancer of fibrosis, both in vivo and in vitro. Furthermore, overexpression of EGR1 increased activation of fibroblasts, while EGR1 knockdown achieved an opposite effect, indicating that EGR1 plays a key role in the inhibitory effect of taurine on TGF-ß-induced fibrosis. CONCLUSIONS: Reduced epidural fibrosis in vivo and decreased activation of fibroblasts in vitro after taurine treatment was mediated by EGR1. Taurine promises to be a potential prevention for epidural fibrosis after laminectomy.

Population Pharmacokinetics of Amitriptyline After Intrathecal, Epidural, and Intravenous Administration in Sheep.

BACKGROUND: Amitriptyline (AMI) is a lipophilic, tricyclic antidepressant with analgesic properties that could potentially be used for epidural (EPI) analgesia. However, no pharmacokinetic data are available for AMI in spinal spaces. The objective of this study was to evaluate the spinal disposition and intrathecal (IT) bioavailability of AMI after IT and EPI administration. METHODS: Six Lacaune ewes received 3 consecutive administrations of AMI. They initially received 10 mg of AMI administered intravenously, then 5 mg of AMI administered intrathecally, and 50 mg of AMI injected into the EPI space. Consecutive administrations were separated by intervals of 2 hours. A simultaneous microdialysis technique was used to determine the EPI and IT concentrations of AMI. Population analysis with S-ADAPT software was used to evaluate the pharmacokinetic parameters. RESULTS: Following intravenous administration, the clearance and central compartment (Vc) in plasma were 1.32 L/min and 147 L, respectively. Concentration-time profiles for the IT and EPI compartments were highly variable after transmeningeal diffusion. The IT Vc after IT administration and the EPI Vc after EPI administration were 2.4 and 48.9 mL, respectively. Less AMI transferred from the EPI to the IT space than from the IT to the EPI compartment, with bioavailabilities of 1.3% and 55%, respectively. CONCLUSIONS: Simultaneous population analysis for AMI demonstrated differences in EPI and IT pharmacokinetics following the EPI and IT administration of this drug. The IT bioavailability of AMI after EPI administration is relatively low.

Clinical application of a novel developed pressure bladder indicator in lumbar epidural puncture.

STUDY OBJECTIVE: A novel pressure bladder indicator was developed, and this study aimed to evaluate the clinical application of the pressure bladder indicator by measuring the epidural space pressure and bladder working pressure on patients undergoing lumbar epidural puncture. DESIGN: Randomized, prospective, double-blinded study PATIENTS: 130 patients SETTING: The Second Hospital of Shandong University INTERVENTIONS: In this study, 60 patients undergoing surgical procedures under lumbar epidural anesthesia were enrolled to detect epidural pressure, and other 70 patients who were undergoing lumbar epidural anesthesia or combined spinal-epidural anesthesia were enrolled to evaluate the pressure bladder indicator. MEASUREMENTS: After successful breakthrough of ligamentum flavum by traditional methods, a pressure transducer was connected to an epidural needle tail and a monitor to measure the epidural pressure at L1-L5 in 60 patients. The working pressure of the bladder was also measured by a transducer. Then lumbar epidural puncture was performed with the pressure bladder indicator in other 70 patients. MAIN RESULTS: The lumbar epidural pressure of the 60 patients was 9.8 ± 4.3 mm Hg, and the bladder working pressure of the pressure bladder indicator was 122 ± 15 mm Hg. All these 70 patients were confirmed with successful bladder indication and lumbar epidural puncture. Thus, the coincidence ratio was 100%. CONCLUSIONS: The novel developed pressure bladder indicator was a reliable and useful technique to conduct successful lumbar epidural puncture.

CCN2 and CCN5 exerts opposing effect on fibroblast proliferation and transdifferentiation induced by TGF-ß.

Epidural fibrosis might occur after lumbar discectomy and contributes to failed back syndrome. Transforming growth factor (TGF)-ß has been reported to influence multiple organ fibrosis, in which connective tissue growth factor/cysteine-rich 61/nephroblastoma overexpressed 2 (CCN2) and CCN5 are involved. However, the effect of CCN2 and CCN5 on TGF-ß induced fibrosis has not yet been elucidated. This study reports that CCN2 and CCN5 play opposing roles in cell proliferation and transdifferentiation of human skin fibroblasts or rabbit epidural scar-derived fibroblasts exposed to TGF-ß. We observed that TGF-ß1 induced fibroblasts proliferation and differentiation in a dose-dependent manner (from 0 µg/L to 20 µg/L). Meanwhile, CCN2 expression is up-regulated while CCN5 expression is inhibited by TGF-ß1 exposure. Furthermore, it is demonstrated that CCN2 overexpression leads to promoted proliferation and elevated collagen and α-smooth muscle actin (α-SMA) expression, which are inhibited by CCN5 overexpression. Moreover, it is shown that the cysteine knot (CT) domain, present in CCN2 but absent in CCN5, plays an essential part in fibroblast proliferation and differentiation. Additionally, enhanced TGF-ß and CCN2 expression but decreased CCN5 expression is found in rabbit epidural scar-derived fibroblasts. Overall, the results show the opposing effects of CCN2 and CCN5 on fibroblast proliferation and transdifferentiation induced by TGF-ß.

Detection of the communicating hole(s) of spinal extradural arachnoid cysts using time-spatial labeling inversion pulse magnetic resonance imaging.

STUDY DESIGN: Report of 2 cases. OBJECTIVE: To report the usefulness of time-spatial labeling inversion pulse magnetic resonance imaging (T-SLIP MRI) for detection of the communicating hole(s) of spinal extradural arachnoid cysts (SEACs). SUMMARY OF BACKGROUND DATA: SEACs normally communicate with the subarachnoid space via small communicating hole(s) in the dura. It is necessary to identify the accurate locations of these communicating hole(s) before attempting to close them through limited laminotomy/laminectomy. Myelocomputed tomography or conventional MRI may fail to detect the locations of the hole(s) because they comprise small dural defects. METHODS: Case 1: A 33-year-old female presented with an SEAC at the T11­L2 vertebral level. Case 2: An 82-year-old female presented with an SEAC at T12­L4 vertebral level. RESULTS: Case 1: T-SLIP MR image of the left parasagittal plane (not the midsagittal or right parasagittal plane) revealed cerebrospinal fluid flow from the subarachnoid space into the cyst at L1. After limited laminotomy at T12­L1 and partial cyst resection, we identified 2 contiguous dural holes immediately medial to the left L1 pedicle; this corroborated the preoperative T-SLIP MRI findings. The holes were sutured. Postoperative conventional MR image confirmed significant cyst shrinkage. Case 2: T-SLIP MR image revealed a curved line at the L1 pedicle in the right parasagittal plane. After L1 laminectomy and partial cyst resection, a dural hole was identified L1 pedicle, which was in agreement with the preoperative T-SLIP MRI findings. After surgery, the lower extremity pain disappeared. Postoperative conventional MR image revealed significant cyst shrinkage. CONCLUSION: T-SLIP MRI is useful for detection of the communicating hole(s) of SEACs. LEVEL OF EVIDENCE: N/A.

Dominance of chemokine ligand 2 and matrix metalloproteinase-2 and -9 and suppression of pro-inflammatory cytokines in the epidural compartment after intervertebral disc extrusion in a canine model.

BACKGROUND CONTEXT: In canine intervertebral disc (IVD) disease, a useful animal model, only little is known about the inflammatory response in the epidural space. PURPOSE: To determine messenger RNA (mRNA) expressions of selected cytokines, chemokines, and matrix metalloproteinases (MMPs) qualitatively and semiquantitatively over the course of the disease and to correlate results to neurologic status and outcome. STUDY DESIGN/SETTING: Prospective study using extruded IVD material of dogs with thoracolumbar IVD extrusion. PATIENT SAMPLE: Seventy affected and 13 control (24 samples) dogs. OUTCOME MEASURES: Duration of neurologic signs, pretreatment, neurologic grade, severity of pain, and outcome were recorded. After diagnostic imaging, decompressive surgery was performed. METHODS: Messenger RNA expressions of interleukin (IL)-1ß, IL-2, IL-4, IL-6, IL-8, IL-10, tumor necrosis factor (TNF), interferon (IFN)γ, MMP-2, MMP-9, chemokine ligand (CCL)2, CCL3, and three housekeeping genes was determined in the collected epidural material by Panomics 2.0 QuantiGene Plex technology. Relative mRNA expression and fold changes were calculated. Relative mRNA expression was correlated statistically to clinical parameters. RESULTS: Fold changes of TNF, IL-1ß, IL-2, IL-4, IL-6, IL-10, IFNγ, and CCL3 were clearly downregulated in all stages of the disease. MMP-9 was downregulated in the acute stage and upregulated in the subacute and chronic phase. Interleukin-8 was upregulated in acute cases. MMP-2 showed mild and CCL2 strong upregulation over the whole course of the disease. In dogs with severe pain, CCL3 and IFNγ were significantly higher compared with dogs without pain (p=.017/.020). Dogs pretreated with nonsteroidal anti-inflammatory drugs revealed significantly lower mRNA expression of IL-8 (p=.017). CONCLUSIONS: The high CCL2 levels and upregulated MMPs combined with downregulated T-cell cytokines and suppressed pro-inflammatory genes in extruded canine disc material indicate that the epidural reaction is dominated by infiltrating monocytes differentiating into macrophages with tissue remodeling functions. These results will help to understand the pathogenic processes representing the basis for novel therapeutic approaches. The canine IVD disease model will be rewarding in this process.

Distribution of methylene blue after injection into the epidural space of anaesthetized pregnant and non-pregnant sheep.

The aim of the study was to determine the distribution of different volumes of methylene blue solution injected into the epidural space in anaesthetized pregnant and non-pregnant sheep, to evaluate its cranial distribution and to compare between them. Fifteen pregnant and fifteen non-pregnant sheep were included in the study. Sheep were anaesthetized and received 0.05, 0.1, or 0.2 mL/kg of a lumbosacral epidural solution containing 0.12% methylene blue in 0.9% saline. Thirty minutes after the epidural injection, the ewes were euthanized. The extension of the dye within the epidural space was measured, and the correlation between the volume of the dye injected and the number of stained vertebrae was evaluated. The cranial migration of the dye between pregnant and non-pregnant sheep was also compared. The results show that the volume of methylene blue injected epidurally into pregnant and non-pregnant sheep correlated directly with its cephalic distribution into the epidural space; and a volume of 0.1 mL/kg or 0.2 mL/kg stained up to the first lumbar segment in pregnant and non-pregnant sheep, respectively. Also, the results suggest that the volume of drugs administered into the epidural space of pregnant sheep should be half the volume that would be used in non-pregnant sheep.