Full-Endoscopic Sacroiliac Joint Denervation for Painful Sacroiliac Joint Dysfunction: A Prospective 2-Year Clinical Outcomes and Predictors for Improved Outcomes.

BACKGROUND AND OBJECTIVES: Full-endoscopic sacroiliac joint denervation (FE-SJD) is a novel technique for the management of pain secondary to sacroiliac joint dysfunction. The aim of this study was to assess the long-term efficacy, safety, clinical outcomes, and outcome predictors of uniportal full-endoscopic sacroiliac joint denervation. METHODS: From 2019 to 2021, a total of 47 consecutive patients with pain secondary to sacroiliac joint dysfunction underwent uniportal FE-SJD through posterior approach by a single fellowship-trained spine surgeon. A retrospective analysis of perioperative parameters, complications, and clinical outcomes were obtained prospectively. RESULTS: The patient cohort had a mean age of 59.4 ± 14.0 years, with 63.8% females. Symptom duration averaged 62.1 ± 53.7 months. The mean operative time was 57.1 ± 16.8 minutes. All patients were discharged on the same day of surgery. Significant improvement was noted in preoperative visual analog score (back) and Oswestry Disability Index scores at 3, 6, 12 months, and 2 years (P < .001). Thirty-four patients (72.3%) returned to normal functioning with an average of 82% pain relief and a satisfaction rate of 78.7% at a mean follow-up of 18.2 ± 13.1 months. There were no intraoperative complications. One patient had postoperative right L5 dysesthesia. Seven patients (14.9%) underwent contralateral FE-SJD due to satisfaction with the index procedure but residual pain on the contralateral side. Concomitant lumbar issues correlated with less functional improvement at 2 years (P = .009). CONCLUSION: The long-term clinical results of FE-SJD are favorable. Endoscopic denervation of the dorsal rami branches supplying the sacroiliac joint represents a safe, effective, and durable option to address pain secondary to sacroiliac joint dysfunction. A significant factor that influences outcomes is the presence of concomitant lumbar pathology. Further research is needed to compare this technique with current available treatment options.

ADAM17 activity and other mechanisms of soluble L-selectin production during death receptor-induced leukocyte apoptosis.

L-selectin is an adhesion molecule expressed by neutrophils that broadly directs their infiltration in to sites of inflammation. It is also present at relatively high levels in the serum of normal individuals. It is well established that L-selectin is efficiently shed from the surface of neutrophils upon their activation, a process that regulates its density and binding activity. Neutrophil programmed cell death is critical for the resolution of inflammation, and L-selectin downregulation is induced during this process as well. The mechanisms underpinning this latter process are much less understood, and were investigated in this study. Using a disintegrin and metalloprotease (ADAM)-17 radiation chimeric mice, we demonstrate for the first time that during early events of death receptor-mediated neutrophil apoptosis, L-selectin downregulation occurs primarily by ADAM17-mediated shedding. This was observed as well upon using shRNA to knock down ADAM17 expression in Jurkat cells, a well-studied cell line in terms of the molecular processes involved in the induction of apoptosis. These findings directly reveal that ADAM17 activity occurs during programmed cell death. Hence, the cleavage of particular ADAM17 substrates may be an additional component of the anti-inflammatory program initiated by apoptotic neutrophils. Of interest was that during later stages of induced leukocyte apoptosis, soluble L-selectin production occurred independent of ADAM17, as well as membrane events, such as blebbing and microparticle production. This process may provide an explanation for the lack of diminished serum L-selectin levels in ADAM17-null mice, and suggests a mechanism for the homeostatic maintenance of soluble L-selectin levels in the blood of healthy individuals.

Regulation of mature ADAM17 by redox agents for L-selectin shedding.

L-selectin is constitutively expressed by neutrophils and plays a key role in directing these cells to sites of inflammation. Upon neutrophil activation, L-selectin is rapidly and efficiently down-regulated from the cell surface by ectodomain shedding. We have directly shown that A disintegrin and metalloprotease 17 (ADAM17) is a primary and nonredundant sheddase of L-selection by activated neutrophils in vivo. Following cell activation, intracellular signals lead to the induction of ADAM17's enzymatic activity; however, the target of this inducer mechanism remains unclear. Our study provides evidence of an activation mechanism that involves the extracellular region of the mature form of cell surface ADAM17 and not its intracellular region. We demonstrate that the catalytic activity of purified ADAM17 lacking a prodomain and its intracellular region is diminished under mild reducing conditions by DTT and enhanced by H(2)O(2) oxidation. Moreover, H(2)O(2) reversed ADAM17 inhibition by DTT. The treatment of neutrophils with H(2)O(2) also induced L-selectin shedding in an ADAM17-dependent manner. These findings suggest that thiol-disulfide conversion occurring in the extracellular region of ADAM17 may be involved in its activation. An analysis of ADAM17 revealed that within its disintegrin/cysteine-rich region are two highly conserved, vicinal cysteine sulfhydryl motifs (cysteine-X-X-cysteine), which are well-characterized targets for thiol-disulfide exchange in various other proteins. Using a cell-based ADAM17 reconstitution assay, we demonstrate that the cysteine-X-X-cysteine motifs are critical for L-selectin cleavage. Taken together, our findings suggest that reduction-oxidation modifications of cysteinyl sulfhydryl groups in mature ADAM17 may serve as a mechanism for regulating the shedding of L-selectin following neutrophil stimulation.

Role of ADAM17 in the ectodomain shedding of TNF-alpha and its receptors by neutrophils and macrophages.

TNF-alpha and its receptors TNFRI and TNFRII are cleaved from the surface of leukocytes by a proteolytic process referred to as ectodomain shedding. The role of a disintegrin and metalloproteinase 17 (ADAM17) in this process by the major professional phagocytes neutrophils and macrophages, the primary producers of TNF-alpha during inflammation induction, is based entirely on indirect evidence, and other sheddases have been implicated as well. As Adam17 gene-targeting in mice is lethal, we assessed the protease's relative contribution to TNF-alpha, TNFRI, and TNFRII shedding using radiation chimeric mice with leukocytes lacking functional ADAM17. We report ablated, soluble TNF-alpha, TNFRI, and TNFRII production by neutrophils and macrophages stimulated with various microbial antigens and greatly reduced TNF-alpha levels in vivo following inflammation induction. This is the first simultaneous analysis of TNF-alpha, TNFRI, and TNFRII shedding by neutrophils and macrophages and the first direct evidence that ADAM17 is a primary and nonredundant sheddase.

A potential role for fructose-2,6-bisphosphate in the stimulation of hepatic glucokinase gene expression.

The effects of fructose-2,6-bisphosphate (F-2,6-P(2)) on hepatic glucokinase (GK) and glucose-6-phosphatase (G-6-Pase) gene expression were investigated in streptozotocin-treated mice, which exhibited undetectable levels of insulin. Hepatic F-2,6-P(2) levels were manipulated by adenovirus-mediated overexpression of 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase. Streptozotocin treatment alone or with infusion of control adenovirus leads to a dramatic decrease in hepatic F-2,6-P(2) content compared with normal nondiabetic mice. This is accompanied by a 14-fold decrease in GK and a 3-fold increase in G-6-Pase protein levels, consistent with a diabetic phenotype. Streptozotocin-treated mice that were infused with adenovirus-overexpressing an engineered 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase with high kinase activity and little bisphosphatase activity showed high levels of hepatic F-2,6-P(2). Surprisingly, these mice had a 13-fold increase in GK protein and a 2-fold decrease in G-6-Pase protein compared with diabetic controls. The restoration of GK is associated with increases in the phosphorylation of Akt upon increasing hepatic F-2,6-P(2) content. Moreover, the changes in levels of F-2,6-P(2) and Akt phosphorylation revealed a pattern similar to that of streptozotocin mice treated with insulin, indicating that increasing hepatic content of F-2,6-P(2) mimics the action of insulin. Because G-6-Pase gene expression was down-regulated only after the restoration of euglycemia, the effect of F-2,6-P(2) was indirect. Also, the lowering of blood glucose by high F-2,6-P(2) was associated with an increase in hepatic nuclear factor 1-alpha protein, a transcription factor involved in G-6-Pase gene expression. In conclusion, F-2,6-P(2) can stimulate hepatic GK gene expression in an insulin-independent manner and can secondarily affect G-6-Pase gene expression by lowering the level of plasma glucose.

In vivo role of leukocyte ADAM17 in the inflammatory and host responses during E. coli-mediated peritonitis.

Inflammation is the body's initial response to infection, which is harmful when excessive, as exemplified in sepsis inflammatory syndromes. Ectodomain shedding by the membrane metalloprotease ADAM17 is an emerging regulator of inflammation, as it directs the activity of various inflammatory modulators. At this time, however, little is known about the in vivo function of ADAM17. Here, we show that ADAM17-deficient leukocytes afforded mice a survival benefit following Escherichia coli-mediated peritoneal sepsis, which was associated with a reduction in systemic proinflammatory cytokine levels and bacterial burden. A more rapid yet transitory neutrophil infiltration into the peritoneal cavity of conditional ADAM17 knockout mice was observed when compared with control mice, suggesting a mechanism for their enhanced clearance of bacteria. Preventing the shedding of L-selectin augments neutrophil recruitment, and we show that L-selectin shedding by peritoneal neutrophils in conditional ADAM17 knockout mice was impaired. Moreover, their peritoneal TNF-alpha levels were markedly lower than control mice following E. coli challenge. These events indicate key molecular processes involved in the altered time course of neutrophil recruitment in conditional ADAM17 knockout mice. Overall, our study provides novel in vivo evidence of the instrumental role of ADAM17 in modulating inflammation and host resistance during Gram-negative bacterial infection.

ADAM17 deficiency by mature neutrophils has differential effects on L-selectin shedding.

L-selectin directs neutrophils to sites of inflammation, and upon their activation, surface expression of the receptor is rapidly down-regulated by ectodomain shedding. Tumor necrosis factor-alpha-converting enzyme (TACE, or ADAM17) is a sheddase of L-selectin; however, Adam17 gene targeting (ADAM17(DeltaZn/DeltaZn)) in mice is perinatal lethal and its role in L-selectin shedding by mature neutrophils has not been determined. This was addressed here by using radiation-chimeric mice reconstituted with ADAM17(DeltaZn/DeltaZn) fetal liver cells. ADAM17-deficient neutrophils, monocytes, and lymphocytes failed to shed L-selectin in response to PMA, as did neutrophils infiltrating the inflamed peritoneum. In addition, the absence of functional ADAM17 resulted in significantly increased levels of L-selectin surface expression by peripheral-blood leukocytes, indicating the sheddase also plays a role in the constitutive cleavage of L-selectin. Interestingly, not all manners of L-selectin turnover required ADAM17. Plasma L-selectin levels were similar between ADAM17(DeltaZn/DeltaZn)-chimeric and control mice, as was the shedding of L-selectin by neutrophils undergoing spontaneous apoptosis. The latter process, however, was diminished by a metalloprotease inhibitor, indicating the role of a sheddase other than ADAM17. Together, our data reveal that L-selectin's surface density on neutrophils is regulated by ADAM17, but homeostatic L-selectin cleavage is not.

ADAM17 activity during human neutrophil activation and apoptosis.

Substrates of the metalloprotease ADAM17 (also known as TNF-alpha converting enzyme or TACE) undergo ectodomain shedding and include various inflammatory modulators. Though polymorphonuclear leukocytes contribute significantly to inflammation, direct analyses of ADAM17 on human neutrophils are very limited. In addition, the current understanding of the processes regulating ADAM17 activity primarily relate to its rapid activation. Therefore, to extend insights into the mechanisms of ADAM17 activity, we examined its surface expression and the shedding of its substrates during extended periods of neutrophil activation and apoptosis. Contrary to studies with immortalized hematopoietic cell lines, we report that surface expression of ADAM17 is maintained by human neutrophils activated with formyl peptides or by FcR/complement receptor-mediated phagocytosis. Interestingly, bacterial phagocytosis resulted in a significant increase in ADAM17 expression several hours after pathogen engulfment. We provide novel evidence that ADAM17 surface expression is also maintained during spontaneous and anti-Fas-induced neutrophil apoptosis. The well-validated ADAM17 substrates L-selectin and proTNF-alpha were shed efficiently by neutrophils under each of the conditions tested. Our data thus indicate prolonged ADAM17 expression during neutrophil effector functions. The implications of this may be a role by ADAM17 in both the induction and down-regulation of neutrophil activity.

N-RAP scaffolds I-Z-I assembly during myofibrillogenesis in cultured chick cardiomyocytes.

N-RAP is a muscle-specific protein with an N-terminal LIM domain (LIM), C-terminal actin-binding super repeats homologous to nebulin (SR) and nebulin-related simple repeats (IB) in between the two. Based on biochemical data, immunofluorescence analysis of cultured embryonic chick cardiomyocytes and the targeting and phenotypic effects of these individual GFP-tagged regions of N-RAP, we proposed a novel model for the initiation of myofibril assembly in which N-RAP organizes alpha-actinin and actin into the premyofibril I-Z-I complexes. We tested the proposed model by expressing deletion mutants of N-RAP (i.e. constructs containing two of the three regions of N-RAP) in chick cardiomyocytes and observing the effects on alpha-actinin and actin organization into mature sarcomeres. Although individually expressing either the LIM, IB, or SR regions of N-RAP inhibited alpha-actinin assembly into Z-lines, expression of either the LIM-IB fusion or the IB-SR fusion permitted normal alpha-actinin organization. In contrast, the LIM-SR fusion (LIM-SR) inhibited alpha-actinin organization into Z-lines, indicating that the IB region is critical for Z-line assembly. While permitting normal Z-line assembly, LIM-IB and IB-SR decreased sarcomeric actin staining intensity; however, the effects of LIM-IB on actin assembly were significantly more severe, as estimated both by morphological assessment and by quantitative measurement of actin staining intensity. In addition, LIM-IB was consistently retained in mature Z-lines, while mature Z-lines without significant IB-SR incorporation were often observed. We conclude that the N-RAP super repeats are essential for organizing actin filaments during myofibril assembly in cultured embryonic chick cardiomyocytes, and that they also play an important role in removal of the N-RAP scaffold from the completed myofibrillar structure. This work strongly supports the N-RAP scaffolding model of premyofibril assembly.

New N-RAP-binding partners alpha-actinin, filamin and Krp1 detected by yeast two-hybrid screening: implications for myofibril assembly.

N-RAP, a muscle-specific protein concentrated at myotendinous junctions in skeletal muscle and intercalated disks in cardiac muscle, has been implicated in myofibril assembly. To discover more about the role of N-RAP in myofibril assembly, we used the yeast two-hybrid system to screen a mouse skeletal muscle cDNA library for proteins capable of binding N-RAP in a eukaryotic cell. From yeast two-hybrid experiments we were able to identify three new N-RAP binding partners: alpha-actinin, filamin-2, and Krp1 (also called sarcosin). In vitro binding assays were used to verify these interactions and to identify the N-RAP domains involved. Three regions of N-RAP were expressed as His-tagged recombinant proteins, including the nebulin-like super repeat region (N-RAP-SR), the N-terminal LIM domain (N-RAP-LIM), and the region of N-RAP in between the super repeat region and the LIM domain (N-RAP-IB). We detected significant alpha-actinin binding to N-RAP-IB and N-RAP-LIM, filamin binding to N-RAP-SR, and Krp1 binding to N-RAP-SR and N-RAP-IB. During myofibril assembly in cultured chick cardiomyocytes, N-RAP and filamin appear to co-localize with alpha-actinin in the earliest myofibril precursors found near the cell periphery, as well as in the nascent myofibrils that form as these structures fuse laterally. In contrast, Krp1 is not localized until late in the assembly process, when it appears at the periphery of myofibrils that appear to be fusing laterally. The results suggest that sequential recruitment of N-RAP binding partners may serve an important role during myofibril assembly.