Extracellular Matrix Remodeling Regulates Glucose Metabolism through TXNIP Destabilization.

The metabolic state of a cell is influenced by cell-extrinsic factors, including nutrient availability and growth factor signaling. Here, we present extracellular matrix (ECM) remodeling as another fundamental node of cell-extrinsic metabolic regulation. Unbiased analysis of glycolytic drivers identified the hyaluronan-mediated motility receptor as being among the most highly correlated with glycolysis in cancer. Confirming a mechanistic link between the ECM component hyaluronan and metabolism, treatment of cells and xenografts with hyaluronidase triggers a robust increase in glycolysis. This is largely achieved through rapid receptor tyrosine kinase-mediated induction of the mRNA decay factor ZFP36, which targets TXNIP transcripts for degradation. Because TXNIP promotes internalization of the glucose transporter GLUT1, its acute decline enriches GLUT1 at the plasma membrane. Functionally, induction of glycolysis by hyaluronidase is required for concomitant acceleration of cell migration. This interconnection between ECM remodeling and metabolism is exhibited in dynamic tissue states, including tumorigenesis and embryogenesis.

Methods for the induction of labor: efficacy and safety.

This review assessed the efficacy and safety of pharmacologic agents (prostaglandins, oxytocin, mifepristone, hyaluronidase, and nitric oxide donors) and mechanical methods (single- and double-balloon catheters, laminaria, membrane stripping, and amniotomy) and those generally considered under the rubric of complementary medicine (castor oil, nipple stimulation, sexual intercourse, herbal medicine, and acupuncture). A substantial body of published reports, including 2 large network meta-analyses, support the safety and efficacy of misoprostol (PGE1) when used for cervical ripening and labor induction. Misoprostol administered vaginally at doses of 50 µg has the highest probability of achieving vaginal delivery within 24 hours. Regardless of dosing, route, and schedule of administration, when used for cervical ripening and labor induction, prostaglandin E2 seems to have similar efficacy in decreasing cesarean delivery rates. Globally, although oxytocin represents the most widely used pharmacologic agent for labor induction, its effectiveness is highly dependent on parity and cervical status. Oxytocin is more effective than expectant management in inducing labor, and the efficacy of oxytocin is enhanced when combined with amniotomy. However, prostaglandins administered vaginally or intracervically are more effective in inducing labor than oxytocin. A single 200-mg oral tablet of mifepristone seems to represent the lowest effective dose for cervical ripening. The bulk of the literature assessing relaxin suggests this agent has limited benefit when used for this indication. Although intracervical injection of hyaluronidase may cause cervical ripening, the need for intracervical administration has limited the use of this agent. Concerning the vaginal administration of nitric oxide donors, including isosorbide mononitrate, isosorbide, nitroglycerin, and sodium nitroprusside, the higher incidence of side effects with these agents has limited their use. A synthetic hygroscopic cervical dilator has been found to be effective for preinduction cervical ripening. Although a pharmacologic agent may be administered after the use of the synthetic hygroscopic dilator, in an attempt to reduce the interval to vaginal delivery, concomitant use of mechanical and pharmacologic methods is being explored. Combining the use of a single-balloon catheter with dinoprostone, misoprostol, or oxytocin enhances the efficacy of these pharmacologic agents in cervical ripening and labor induction. The efficacy of single- and double-balloon catheters in cervical ripening and labor induction seems similar. To date, the combination of misoprostol with an intracervical catheter seems to be the best approach when balancing delivery times with safety. Although complementary methods are occasionally used by patients, given the lack of data documenting their efficacy and safety, these methods are rarely used in hospital settings.

Dual Stromal Targeting Sensitizes Pancreatic Adenocarcinoma for Anti-Programmed Cell Death Protein 1 Therapy.

BACKGROUND & AIMS: The stroma in pancreatic ductal adenocarcinoma (PDAC) contributes to its immunosuppressive nature and therapeutic resistance. Herein we sought to modify signaling and enhance immunotherapy efficacy by targeting multiple stromal components through both intracellular and extracellular mechanisms. METHODS: A murine liver metastasis syngeneic model of PDAC was treated with focal adhesion kinase inhibitor (FAKi), anti-programmed cell death protein 1 (PD-1) antibody, and stromal hyaluronan (HA) degradation by PEGylated recombinant human hyaluronidase (PEGPH20) to assess immune and stromal modulating effects of these agents and their combinations. RESULTS: The results showed that HA degradation by PEGPH20 and reduction in phosphorylated FAK expression by FAKi leads to improved survival in PDAC-bearing mice treated with anti-PD-1 antibody. HA degradation in combination with FAKi and anti-PD-1 antibody increases T-cell infiltration and alters T-cell phenotype toward effector memory T cells. FAKi alters the expression of T-cell modulating cytokines and leads to changes in T-cell metabolism and increases in effector T-cell signatures. HA degradation in combination with anti-PD-1 antibody and FAKi treatments reduces granulocytes, including granulocytic- myeloid-derived suppressor cells and decreases C-X-C chemokine receptor type 4 (CXCR4)-expressing myeloid cells, particularly the CXCR4-expressing granulocytes. Anti-CXCR4 antibody combined with FAKi and anti-PD-1 antibody significantly decreases metastatic rates in the PDAC liver metastasis model. CONCLUSIONS: This represents the first preclinical study to identify synergistic effects of targeting both intracellular and extracellular components within the PDAC stroma and supports testing anti-CXCR4 antibody in combination with FAKi as a PDAC treatment strategy.

Acute loading has minor influence on human articular cartilage gene expression and glycosaminoglycan composition in late-stage knee osteoarthritis: a randomised controlled trial.

OBJECTIVE: Osteoarthritis (OA) remains clinically challenging. Regular physical exercise improves symptoms though it is unclear whether exercise influences cartilage at the molecular level. Thus, we aimed to determine the effect of acute loading on gene expression and glycosaminoglycan (GAG) content in human OA cartilage. DESIGN: Patients with primary knee OA participated in this single-blind randomised controlled trial initiated 3.5 h prior to scheduled joint replacement surgery with or without loading by performing one bout of resistance exercise (one-legged leg press). Cartilage from the medial tibia condyle was sampled centrally, under the meniscus, and from peripheral osteophytes. Samples were analysed for gene expression by real-time reverse transcriptase polymerase chain reaction, and hyaluronidase-extracted matrix was analysed for GAG composition by immuno- and dimethyl-methylene blue assays. RESULTS: Of 32 patients randomised, 31 completed the intervention: mean age 69 ± 7.5 years (SD), 58% female, BMI 29.4 ± 4.4 kg/m2. Exercise increased chondroitin sulphate extractability [95% CI: 1.01 to 2.46; P = 0.0486] but cartilage relevant gene expression was unchanged. Regionally, the submeniscal area showed higher MMP-3, MMP-13, IGF-1Ea, and CTGF, together with lower lubricin and COMP expression compared to the central condylar region. Further, osteophyte expression of MMP-1, MMP-13, IGF-1Ea, and TGF-ß3 was higher than articular cartilage and lower for aggrecan, COMP, and FGF-2. Hyaluronidase-extracted matrix from central condylar cartilage contained more GAGs but less chondroitin sulphate compared to submeniscal cartilage. CONCLUSION: Acute exercise had minor influence on cartilage GAG dynamics, indicating that osteoarthritic cartilage is not significantly affected by acute exercise. However, the regional differences suggest a chronic mechanical influence on human cartilage. GOV REGISTRATION NUMBER: NCT03410745.

[Slowdown of replicative aging of fibroblasts by hyaluronan complexes with gold nanoparticles.]

Determination the activity of the genes of sirtuin-1, hyaluronidase, TGF-ß cytokine, calreticulin in the process of replicative aging of human fibroblasts in vitro and the effect of hyaluronan preparations with gold nanoparticles on the activity of replicative cell aging. Compared the expression of proteins of the studied genes using specific markers at 7 and 14 passages of cultivation of fibroblasts isolated from human skin, without drugs and in the presence of drugs in the growth medium. This work shows a decrease in the activity of the sirtuin 1 gene and an increase in the expression of hyaluronidase in the process of replicative aging of human fibroblasts. Found a means of slowing down replicative aging by activating the SIRT-1 gene and reducing the activity of hyaluronidase in action in the growth medium of hyaluronan preparations with gold nanoparticles. The discussed variants of cell transitions to the pathological state, caused by replicative aging and the mechanisms of slowing down the replicative aging of human fibroblasts.

Treating Sunken Upper Eyelid With Hyaluronic Acid: Recommendations and Results.

BACKGROUND: Supraorbital hollowness is a feature that gives an unesthetic, aged, and cadaveric aspect to the eyelids. This complex anatomic area is affected by different variables. Treating this area with fillers poses a real challenge in terms of reducing risks and predicting natural results. We aimed to report outcomes observed in patients treated with injections of hyaluronic acid (HA) soft tissue fillers. Additionally, we have reviewed sunken upper eyelid (SUE) anatomy, etiology, and pathophysiology with the objectives of describing a new classification of SUE, showing a lower risk technique to fill the area, and explaining the importance of using a high cohesivity HA product. METHODS: We included 32 adults (96.9% female) without previous fillers in the area, who were injected with HA (Belotero Balance) with a 25 G, 40 mm blunt cannula. The procedure was fractionated in 2/3 visits. The assessment was performed at day 0, day 14, day 30, and day 365 with standard camera and Vectra H2 (Canfield). RESULTS: All patients treated with HA showed natural results without significant edema on day 30. Only 1 patient (3.1%) required hyaluronidase injections to dissolve overcorrection of the area on day 14. No severe complications were observed. For most patients, obtained results remained stable on day 365. CONCLUSION: As SUE is one of the most prominent signs of aging, the desire for rejuvenation and the popularity of non-surgical solutions have increased. We present our results with a cohesive HA as a highly suitable filler for SUE. Given the high patient satisfaction, long-lasting results, and reduced complication risk, our presented approach may represent a safe and effective novel treatment strategy.J Drugs Dermatol. 2022;21(9):1002-1008. doi:10.36849/JDD.6745R1.

A Connecting Link between Hyaluronan Synthase 3-Mediated Hyaluronan Production and Epidermal Function.

Hyaluronan (HA), an essential component of the extracellular matrix of the skin, is synthesized by HA synthases (HAS1-3). To date, epidermal HA has been considered a major player in regulating cell proliferation and differentiation. However, a previous study reported that depletion of epidermal HA by Streptomyces hyaluronidase (St-HAase) has no influence on epidermal structure and function. In the present study, to further explore roles of epidermal HA, we examined effects of siRNA-mediated knockdown of HAS3, as well as conventional HA-depletion methods using St-HAase and 4-methylumbelliferone (4MU), on epidermal turnover and architecture in reconstructed skin or epidermal equivalents. Consistent with previous findings, HA depletion by St-HAase did not have a substantial influence on the epidermal architecture and turnover in skin equivalents. 4MU treatment resulted in reduced keratinocyte proliferation and epidermal thinning but did not seem to substantially decrease the abundance of extracellular HA. In contrast, siRNA-mediated knockdown of HAS3 in epidermal equivalents resulted in a significant reduction in epidermal HA content and thickness, accompanied by decreased keratinocyte proliferation and differentiation. These results suggest that HAS3-mediated HA production, rather than extracellularly deposited HA, may play a role in keratinocyte proliferation and differentiation, at least in the developing epidermis in reconstructed epidermal equivalents.

The Bovhyaluronidase Azoximer (Longidaza®) Disrupts Candida albicans and Candida albicans-Bacterial Mixed Biofilms and Increases the Efficacy of Antifungals.

Background and Objectives: Candida albicans causes various diseases ranging from superficial mycoses to life-threatening systemic infections often associated with biofilm formation, including mixed fungal−bacterial consortia. The biofilm matrix protects cells, making Candida extremely resistant to treatment. Here, we show that the bovhyaluronidase azoximer (Longidaza®) in vitro destroys the biofilm formed by either C. albicans alone or mixed with bacteria, this way decreasing the concentrations of antimicrobials required for the pathogen's eradication. Materials and Methods: Bovhyaluronidase azoximer, Longidaza® was obtained from NPO Petrovax Pharm Ltd., Moscow, Russia as lyophilized powder. The antifungal activity was assessed by microdilution assay and CFUs counting. Antibiofilm activity was evaluated via biofilms staining and scanning electron microscopy. Results: Thus, treatment with Longidaza® reduced the biofilm biomass of nine C. albicans clinical isolates by 30−60%, while mixed biofilms of C. albicans with various bacteria were destroyed by 30−40%. Furthermore, the concentration of fluconazole required to achieve a similar reduction of the residual respiratory activity of detached cell clumps of four C. albicans isolates has been reduced four-fold when combined with Longidaza®. While in the biofilm, two of four isolates became significantly more susceptible to fluconazole in combination with Longidaza®. Conclusion: Taken together, our data indicate that Longidaza® is capable of suppression of tissues and artificial surfaces biofouling by C. albicans biofilms, as well as facilitating drug penetration into the cell clumps, this way decreasing the effective MIC of antifungals.

Postpartum Involution of Mouse Myometrium in Acute CCl4-Induced Hepatosis and Its Correction with Immobilized Hyaluronidase.

We studied structural rearrangement of the myometrium of C57BL/6 mice during the postpartum period under conditions of acute toxic hepatosis induced by CCl4 injection and its correction with immobilized hyaluronidase. In contrast to physiological pregnancy, involution of the myometrium in mice under conditions of acute toxic hepatosis were not completed by the 10th day of the postpartum period. When toxic hepatosis was corrected with immobilized hyaluronidase, the main mechanism of postpartum involution of the mouse myometrium was clasmacytosis, the process of postpartum involution was significantly accelerated, but not completed by the 10th day, probably due to reduced vascularization of the myometrium.

Micellar Hyaluronidase and Spiperone as a Potential Treatment for Pulmonary Fibrosis.

Concentration of hyaluronic acid (HA) in the lungs increases in idiopathic pulmonary fibrosis (IPF). HA is involved in the organization of fibrin, fibronectin, and collagen. HA has been proposed to be a biomarker of fibrosis and a potential target for antifibrotic therapy. Hyaluronidase (HD) breaks down HA into fragments, but is a subject of rapid hydrolysis. A conjugate of poloxamer hyaluronidase (pHD) was prepared using protein immobilization with ionizing radiation. In a model of bleomycin-induced pulmonary fibrosis, pHD decreased the level of tissue IL-1ß and TGF-ß, prevented the infiltration of the lung parenchyma by CD16+ cells, and reduced perivascular and peribronchial inflammation. Simultaneously, a decrease in the concentrations of HA, hydroxyproline, collagen 1, total soluble collagen, and the area of connective tissue in the lungs was observed. The effects of pHD were significantly stronger compared to native HD which can be attributed to the higher stability of pHD. Additional spiperone administration increased the anti-inflammatory and antifibrotic effects of pHD and accelerated the regeneration of the damaged lung. The potentiating effects of spiperone can be explained by the disruption of the dopamine-induced mobilization and migration of fibroblast progenitor cells into the lungs and differentiation of lung mesenchymal stem cells (MSC) into cells of stromal lines. Thus, a combination of pHD and spiperone may represent a promising approach for the treatment of IPF and lung regeneration.

Immunomodulatory activity of hyaluronidase is associated with metabolic adaptations during acute inflammation.

OBJECTIVE AND DESIGN: Investigate survival outcomes, and immunological and metabolomic effects of hyaluronidase (Hz) treatment during mouse models of acute inflammation and sepsis. METHODS: Survival of C57Bl/6 mice was monitored after lethal challenge with lipopolysaccharide (LPS) or cecal and ligation puncture (CLP)-induced sepsis and treated with Hz or saline. Mice were also challenged with LPS and treated with Hz for leukocyte counting, cytokine quantification and determination of metabolomic profiles in the peritoneal fluid. RESULTS: Hz treatment improved survival outcomes after lethal challenge with LPS or CLP-induced sepsis. LPS challenge promoted acute neutrophil accumulation and production of interleukin-1ß (IL-1ß) and IL-6 in the peritoneum, whereas Hz treatment suppressed neutrophil infiltration and cytokine production. We further characterized the metabolomic alterations caused by LPS challenge, which predicted activity of metabolic pathways related to fatty acids and eicosanoids. Hz treatment had a profound effect over the metabolic response, reflected by reductions of the relative levels of fatty acids. CONCLUSION: Collectively, these data demonstrate that Hz treatment is associated with metabolic reprogramming of pathways that sustain the inflammatory response.

Hyaluronidase in Dermatology: Uses Beyond Hyaluronic Acid Fillers.

Hyaluronidase is mostly widely recognized for its off-label use in correction of complications of hyaluronic acid fillers. However, its utility in other aspects of dermatology is less widely acknowledged. We describe the varied uses of hyaluronidase in dermatology and the underlying evidence base for its dermatological indications. This includes its uses in enhancing drug delivery (for local anesthesia, keloid and hypertrophic scars, and for Kaposi’s sarcoma), in the treatment of disorders associated with mucin deposition (myxedema, scleroderma, scleredema, and cutis verticis gyrata) and its potential uses in surgery (as a pre-operative adjuvant in dermatofibrosarcoma protuberans, for periorbital edema, and for hematomas). In select circumstances, hyaluronidase might be more efficacious than more established treatments with fewer adverse effects. We propose hyaluronidase as the latest addition to our global dermatological armamentarium and implore dermatologists to consider its use to enhance their practice. J Drugs Dermatol. 2020;19(10):993-998. doi:10.36849/JDD.2020.5416.

CD44 contributes to hyaluronan-mediated insulin resistance in skeletal muscle of high-fat-fed C57BL/6 mice.

Extracellular matrix hyaluronan is increased in skeletal muscle of high-fat-fed insulin-resistant mice, and reduction of hyaluronan by PEGPH20 hyaluronidase ameliorates diet-induced insulin resistance (IR). CD44, the main hyaluronan receptor, is positively correlated with type 2 diabetes. This study determines the role of CD44 in skeletal muscle IR. Global CD44-deficient (cd44-/-) mice and wild-type littermates (cd44+/+) were fed a chow diet or 60% high-fat diet for 16 wk. High-fat-fed cd44-/- mice were also treated with PEGPH20 to evaluate its CD44-dependent action. Insulin sensitivity was measured by hyperinsulinemic-euglycemic clamp (ICv). High-fat feeding increased muscle CD44 protein expression. In the absence of differences in body weight and composition, despite lower clamp insulin during ICv, the cd44-/- mice had sustained glucose infusion rate (GIR) regardless of diet. High-fat diet-induced muscle IR as evidenced by decreased muscle glucose uptake (Rg) was exhibited in cd44+/+ mice but absent in cd44-/- mice. Moreover, gastrocnemius Rg remained unchanged between genotypes on chow diet but was increased in high-fat-fed cd44-/- compared with cd44+/+ when normalized to clamp insulin concentrations. Ameliorated muscle IR in high-fat-fed cd44-/- mice was associated with increased vascularization. In contrast to previously observed increases in wild-type mice, PEGPH20 treatment in high-fat-fed cd44-/- mice did not change GIR or muscle Rg during ICv, suggesting a CD44-dependent action. In conclusion, genetic CD44 deletion improves muscle IR, and the beneficial effects of PEGPH20 are CD44-dependent. These results suggest a critical role of CD44 in promoting hyaluronan-mediated muscle IR, therefore representing a potential therapeutic target for diabetes.

Investigating the role of CD44 and hyaluronate in embryo-epithelial interaction using an in vitro model.

Implantation failure is an important impediment to increasing success rates in assisted reproductive technologies. Knowledge of the cascade of morphological and molecular events at implantation remains limited. Cell surface CD44 and hyaluronate (HA) have been reported in the uterus, but a role in intercellular interaction at implantation remains to be evaluated. Mouse embryos were co-cultured with human Ishikawa endometrial epithelial monolayers over 2 days. Attachment was tenuous during the first 24 h, after which it became stable, leading to breaching of the monolayer. The effects of enzymatically reducing the density of HA, or introducing a function-blocking antibody to CD44, were monitored during progression from weak to stable embryonic attachment. Hyaluronidase-mediated removal of surface HA from the epithelial cells enhanced the speed of attachment, while a similar treatment of embryos had no effect. The antibody to CD44 caused retardation of initial attachment. These results suggest that CD44-HA binding could be employed by embryos during initial docking, but the persistence of HA in epithelial cells might be detrimental to later stages of implantation by retarding attainment of stable attachment.

Effect of diabetes and hyaluronidase on the retinal endothelial glycocalyx in mice.

We sought to investigate the effects of diabetes and hyaluronidase on the thickness of the endothelial glycocalyx layer in the mouse retina. In our study, the retinal circulation of diabetic Ins2(Akita) mice and their nondiabetic littermates were observed via intravital microscopy. The endothelial glycocalyx thickness was determined from the infusion of two fluorescently labeled plasma markers, one of which was a high molecular weight rhodamine dextran (MW = 155,000) excluded from the glycocalyx, and the other a more permeable low molecular weight sodium fluorescein (MW = 376). In nondiabetic C57BL/6 mice, the glycocalyx thickness also was evaluated prior to and following infusion of hyaluronidase, an enzyme that can degrade hyaluronic acid on the endothelial surface. A leakage index was used to evaluate the influence of hyaluronidase on the transport of the fluorescent tracers from the plasma into the surrounding tissue, and plasma samples were obtained to measure levels of circulating hyaluronic acid. Both diabetes and hyaluronidase infusion significantly reduced the thickness of the glycocalyx in retinal arterioles (but not in venules), and hyaluronidase increased retinal microvascular leakage of both fluorescent tracers into the surrounding tissue. However, only hyaluronidase infusion (not diabetes) increased circulating plasma levels of hyaluronic acid. In summary, our findings demonstrate that diabetes and hyaluronidase reduce the thickness of the retinal endothelial glycocalyx, in which hyaluronic acid may play a significant role in barrier function.

Clinical Applications of Hyaluronidase.

Hyaluronidases are enzymes that degrade hyaluronic acid, which constitutes an essential part of the extracellular matrix. Initially discovered in bacteria, hyaluronidases are known to be widely distributed in nature and have been found in many classes including insects, snakes, fish and mammals. In the human, six different hyaluronidases, HYAL1-4, HYAL-P1 and PH-20, have been identified. PH-20 exerts the strongest biologic activity, is found in high concentrations in the testicles and can be localized on the head and the acrosome of human spermatozoa. Today, animal-derived bovine or ovine testicular hyaluronidases as well as synthetic hyaluronidases are clinically applied as adjuncts to increase the bioavailability of drugs, for the therapy of extravasations, or for the management of complications associated with the aesthetic injection of hyaluronic acid-based fillers. Further applications in the fields of surgery, aesthetic medicine, immunology, oncology, and many others can be expected for years to come. Here, we give an overview over the molecular and cellular mode of action of hyaluronidase and the hyaluronic acid metabolism, as well as over current and potential future clinical applications of hyaluronidase.

Hyaluronidase Pretreatment Improves the Cryopreservation of Human Ovarian Tissue.

BACKGROUND: Slow freezing has been widely applied to preserve human ovarian tissue. OBJECTIVE: The aim of this study is to determine whether human ovarian cortex pretreated with hyaluronidase can improve the protective effect of cryopreservation. MATERIALS AND METHODS: Human ovarian biopsies from seven patients were cryopreserved using slow freezing. The samples in the experimental group were incubated in culture medium with hyaluronidase for a short time before cryopreservation. The histology of the thawed ovarian tissue was investigated; the levels of steroid hormone in the culture media were then used to evaluate the development and function of thawed ovarian tissue. RESULTS: The result showed that the percentage of morphologically abnormal primordial follicles was higher in the freezing control than in the experimental group (P<0.05). 17ß-estradiol (E2) and progesterone (P4) concentrations were significantly higher in the experimental group than in the freezing control group (P<0.05). CONCLUSION: Our study showed that human ovarian cortex pretreatment with hyaluronidase can improve the protective effect of cryopreservation by improving the penetration of cryoprotectant.

Effects of hyaluronidase on ropivacaine or bupivacaine regional anaesthesia of the canine pelvic limb.

OBJECTIVE: To determine the effect of hyaluronidase on time to onset and offset of anaesthesia in ropivacaine or bupivacaine femoral-ischiatic nerve blocks. STUDY DESIGN: Blinded randomized crossover trial. ANIMALS: Eight dogs. METHODS: Each dog underwent four treatments separated into two blocks - initially, the ropivacaine treatment block: RS (ropivacaine 0.5% plus saline 0.9%) and RH (ropivacaine 0.5% plus hyaluronidase 100 IU mL-1), followed 3 weeks later by the bupivacaine treatment block: BS (bupivacaine 0.5% plus saline) and BH (bupivacaine 0.5% plus hyaluronidase). The local anaesthetics were administered at 0.1 mL kg-1 per site. Hyaluronidase and saline were administered at 0.02 mL kg-1 per site. Performance of femoral-ischiatic blocks was aided by a combined ultrasound-electrolocation technique. The mechanical nociceptive threshold was measured, until offset or 360 minutes, using an algometer to ascertain baseline, onset and offset of anaesthesia. Onset and offset of anaesthesia were defined as a 25% increase above and as a return to <25% above baseline nociceptive threshold readings, respectively. RESULTS: The median (range) onset of anaesthesia for RS and RH was 21 (3-60) and 12 (3-21) minutes, respectively (p = 0.141), and offset was 270 (90-360) and 180 (30-300) minutes, respectively (p = 0.361). By contrast, the median (range) onset of anaesthesia for BS and BH was 24 (3-60) and 9 (3-27) minutes, respectively (p = 0.394), and offset was 360 (240-360) and 330 (210-360) minutes, respectively (p = 0.456). CONCLUSION AND CLINICAL RELEVANCE: Hyaluronidase had no effect on the onset and offset times of ropivacaine and bupivacaine femoral-ischiatic nerve blocks in dogs compared with saline. The onset and offset times were highly variable in all treatments. Clinically, the high variability of the onset and offset times of the regional anaesthesia of these local anaesthetic drugs means that clinicians must monitor the animal's response and, if required, provide additional analgesic drugs.

Mechanisms of NMDA Receptor- and Voltage-Gated L-Type Calcium Channel-Dependent Hippocampal LTP Critically Rely on Proteolysis That Is Mediated by Distinct Metalloproteinases.

Long-term potentiation (LTP) is widely perceived as a memory substrate and in the hippocampal CA3-CA1 pathway, distinct forms of LTP depend on NMDA receptors (nmdaLTP) or L-type voltage-gated calcium channels (vdccLTP). LTP is also known to be effectively regulated by extracellular proteolysis that is mediated by various enzymes. Herein, we investigated whether in mice hippocampal slices these distinct forms of LTP are specifically regulated by different metalloproteinases (MMPs). We found that MMP-3 inhibition or knock-out impaired late-phase LTP in the CA3-CA1 pathway. Interestingly, late-phase LTP was also decreased by MMP-9 blockade. When both MMP-3 and MMP-9 were inhibited, both early- and late-phase LTP was impaired. Using immunoblotting, in situ zymography, and immunofluorescence, we found that LTP induction was associated with an increase in MMP-3 expression and activity in CA1 stratum radiatum. MMP-3 inhibition and knock-out prevented the induction of vdccLTP, with no effect on nmdaLTP. L-type channel-dependent LTP is known to be impaired by hyaluronic acid digestion. We found that slice treatment with hyaluronidase occluded the effect of MMP-3 blockade on LTP, further confirming a critical role for MMP-3 in this form of LTP. In contrast to the CA3-CA1 pathway, LTP in the mossy fiber-CA3 projection did not depend on MMP-3, indicating the pathway specificity of the actions of MMPs. Overall, our study indicates that the activation of perisynaptic MMP-3 supports L-type channel-dependent LTP in the CA1 region, whereas nmdaLTP depends solely on MMP-9. SIGNIFICANCE STATEMENT: Various types of long-term potentiation (LTP) are correlated with distinct phases of memory formation and retrieval, but the underlying molecular signaling pathways remain poorly understood. Extracellular proteases have emerged as key players in neuroplasticity phenomena. The present study found that L-type calcium channel-dependent LTP in the CA3-CA1 hippocampal projection is critically regulated by the activity of matrix metalloprotease 3 (MMP-3), in contrast to NMDAR-dependent LTP regulated by MMP-9. Moreover, the induction of LTP was associated with an increase in MMP-3 expression and activity. Finally, we found that the digestion of hyaluronan, a principal extracellular matrix component, disrupted the MMP-3-dependent component of LTP. These results indicate that distinct MMPs might act as molecular switches for specific types of LTP.

Durability, Behavior, and Tolerability of 5 Hyaluronidase Products.

BACKGROUND: Hyaluronic acid (HA) dermal fillers are commonly used in cosmetic dermatology. Due to differences in their physical characteristics, HA fillers demonstrate different sensitivity to degradation by hyaluronidase (Hase) because of HA concentration and differences in cross-linking. Similarly, there are differences in the activity of Hase products depending on source and concentration. OBJECTIVE: The primary objective was to demonstrate the differences in potency and activity of 5 Hase products when used to degrade 5 different HA products using a human in vivo model. MATERIALS AND METHODS: The study subject was a healthy, consenting adult woman scheduled to undergo abdominoplasty. Skin to be excised was injected with 0.1 to 0.2 mL of each filler (10 injections each) leaving a visible lump. Immediately afterward, the HA lumps were injected with 4 IU of each Hase product every 2 minutes until the HA lumps were no longer visible or palpable. This procedure was repeated after 30 days. Injected tissues were excised after abdominoplasty for histological analysis. RESULTS: The 5 Hase products displayed a wide range of doses and times required to completely degrade the 5 HA products ranging from <2 to >16 minutes. CONCLUSION: Cosmetic practitioners should familiarize themselves with differences in HA and Hase products.