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Background: The collagenase of the bacterium Clostridium histolyticum (CCH) is already an established treatment for fibroproliferative diseases like M. Dupuytren and M. Peyronie Although results are comparable to surgical intervention, skin laceration is a severe and relevant side effect. Doxycycline (DOX) recently rose interest as an inhibitor of matrix-metalloproteinases alongside its capabilities of skin accumulation. It therefore might be a potential skin protective agent in the use of CCH., Methods: For simulation of a fibroproliferative disease adjacent to the skin, we utilized a rodent model of capsular fibrosis involving silicone implants and subsequent fibrotic capsule formation. For in-vitro studies, fibrotic capsules were excised and incubated with 0.9 mg/ml CCH and four different doses of DOX. For in-vivo experiments, animals received 0.0, 0.3 or 0.9 mg/ml CCH injections into the fibrotic capsules with or without prior oral DOX administration. Outcome analysis included histology, immunohistochemistry, gene expression analysis, chemical collagen and DOX concentration measurements as well as μCT imaging., Results: In-vitro, DOX showed a dose-dependent inhibition of CCH activity associated with increasing capsule thickness and collagen density and content. In-vivo, oral DOX administration did neither interfere with capsule formation nor in effectiveness of CCH dissolving fibrotic capsule tissue. However, skin thickness and especially collagen density was significantly higher compared to control groups. This led to a reduced rate of clinical skin lacerations after DOX administration., Conclusion: DOX inhibits CCH and accumulates in the skin. Thereby, DOX can effectively reduce skin laceration after CCH treatment., Competing Interests: The authors have declared that no competing interests exist.

Background: Uterine fibroids are highly prevalent, collagen-rich, mechanically stiff, fibrotic tumors for which new therapeutic options are needed. Increased extracellular matrix (ECM) stiffness activates mechanical signaling and Hippo/YAP promoting fibroid growth, but no prior studies have tested either as a therapeutic target. We tested the hypothesis that injection of a purified form of collagenase Clostridium histolyticum (CCH) that selectively digests type I and type III collagens would alter ECM stiffness, Hippo signaling, and selectively reduce fibroid cell growth. We also used two FDA-approved drugs, verteporfin and nintedanib, to elucidate the role of Hippo/YAP signaling in uterine fibroid and myometrial cells., Methods: The clinical trial was registered (NCT02889848). Stiffness of samples was measured by rheometry. Protein expression in surgical samples was analyzed via immunofluorescence. Protein and gene expression in uterine fibroid or myometrial cell lines were measured by real time PCR and western blot, and immunofluorescence., Results: Injection of CCH at high doses (0.1-0.2 mg/cm 3 ) into fibroids resulted in a 46% reduction in stiffness in injected fibroids compared to controls after 60 days. Levels of the cell proliferation marker proliferative cell nuclear antigen (PCNA) were decreased in fibroids 60 days after injection at high doses of CCH. Key Hippo signaling factors, specifically the transcriptionally inactive phosphorylated YAP (p-YAP), was increased at high CCH doses, supporting the role of YAP in fibroid growth. Furthermore, inhibition of YAP via verteporfin (YAP inhibitor) decreased cell proliferation, gene and protein expression of key factors promoting fibrosis and mechanotransduction in fibroid cells. Additionally, the anti-fibrotic drug, nintedanib, inhibited YAP and showed anti-fibrotic effects., Conclusions: This is the first report that in vivo injection of collagenase into uterine fibroids led to a reduction in Hippo/YAP signaling and crucial genes and pathways involved in fibroid growth. These results indicate that targeting ECM stiffness and Hippo signaling might be an effective strategy for uterine fibroids., (© 2021 The Authors. Clinical and Translational Medicine published by John Wiley & Sons Australia, Ltd on behalf of Shanghai Institute of Clinical Bioinformatics.)

In the present study we aimed to investigate the surgical outcomes of patients with persistent penile curvature (PC) after Collagenase Clostridium histolyticum (CCH) intraplaque injections. Data from 90 patients with persistent PC after CCH in a multicentre study from 6 andrological centres were retrospectively reviewed. Three standardized surgical techniques were performed. Group 1: plaque incision grafting (PIG) with penile prosthesis implant (PPI); Group 2: PIG without PPI; Group 3: Nesbit technique. Hospital stay, operative time, postoperative complications and PC persistency/recurrence (> 20°) were evaluated. Overall satisfaction and functional outcomes were assessed through International Index of Erectile Function-Erectile Function (IIEF-EF), Peyronie's Disease Questionnaire (PDQ), Female Sexual Function Index (FSFI) administered pre and 3 months postoperatively. Of all, 25 (27.8%) patients received grafting procedure + PPI (Group 1), 18 (20.0%) patients belonged to Group 2, and 47 (52.2%) to Group 3. Bovine pericardium graft and collagen fleece have been used in in 22 (51.2%) and 21 (48.8%) patients, respectively. Median penile length after surgery was 13.0 cm (IQR 12.0-15.0). After surgery, Group 1 showed higher increase in penile length after surgery and better improvements in terms of PDQ-PS. In contrast, both IIEF-EF and FSFI scores did not differ among groups. Overall, 86 (95.6%) did not report any complication. 4 (4.4%) patients had PC recurrence; of those, 2 (8.0%), 1 (5.6%) and 1 (2.1%) cases were observed in Group 1, Group 2 and Group 3, respectively. In case of persistent PC after CCH, surgical correction by grafting with or without concomitant PPI or Nesbit technique emerged as a technically feasible, effective and safe procedure, with no significant postoperative complications.

Background: Collagenase Clostridium histolyticum (CCH) is a recent treatment for Dupuytren disease, which is a fibroproliferative disorder that leads to progressive, persistent digital flexion contracture that interferes with basic daily activities. While CCH has changed the treatment of this hand disorder, numerous concerns have to be analyzed., Aims: Our purpose is to assess the current status of this medical treatment., Methods: Literary review based on a manual search on PubMed, Web of Science, and Google Academic., Results: Pharmacoeconomic analyses support the use of CCH, but long-term studies showing that it should be favored over conventional surgery or other treatments are lacking. Treatment decisions, therefore, must be guided by current data, which include a 5-year recurrence rate of 47%. Complications following CCH treatment are also a controversial topic, as rates of over 90% have been reported, although most of the complications are mild and self-limiting. A definition and classification of CCH-related complications is sorely needed. If we exclude adverse effects that could be considered inherent to the mechanisms of action of CCH, then the complication rate would be similar to rates reported for other techniques. Although CCH is becoming an increasingly popular treatment for Dupuytren disease, the potential applications of this modality, are much higher than currently believed, for more disorders characterized by excessive fibrosis., Conclusion: Currently, the administration of this treatment is promising although long-term studies are necessary to see the real role that this drug can play in both Dupuytren's disease and other fibrotic disorders.

Objective: We investigated the expression pattern of long noncoding RNAs (lncRNA) and messenger RNAs (mRNA) from two different intracerebral hemorrhage (ICH) rat models, and performed gene ontology and gene/protein interaction analyses., Methods: We harvested hemorrhagic brain 1, 3, and 7 days after ICH induction by stereotactic collagenase injection. We performed microarray analyses with Agilent array platform to compare the expression of lncRNA and mRNAs from hemorrhagic and normal brains. The RNA expression patterns were also examined from the autologous blood injection ICH model at days 1 and 3, and significantly altered lncRNAs from two ICH models were validated by quantitative reverse transcriptase-polymerase chain reaction. Gene ontology analysis and pathway analysis were performed with differentially expressed mRNAs after ICH. Gene and protein interaction analysis was performed to elucidate the functional role of upregulated lncRNA in neuronal damage., Results: Among the 13,661 lncRNAs studied, 83, 289, and 401 lncRNAs were significantly elevated after 1, 3, and 7 days after collagenase-induced ICH, respectively. NR_027324, or H19, was the most upregulated lncRNA after 1 day from the two ICH models and its elevation persisted until the 7th day. Gene ontology analysis revealed that immune-related biological processes such as immune response, immune system process, and defense response were upregulated from both ICH models. Gene and protein interaction study demonstrated that NR_027324 was closely related to the type I interferon signaling pathway., Interpretation: This study illustrates the dynamic expression pattern of the lncRNA profile following ICH, and that H19 is the most consistently upregulated lncRNA after ICH., (© 2019 The Authors. Annals of Clinical and Translational Neurology published by Wiley Periodicals, Inc on behalf of American Neurological Association.)

Disruption of the blood-brain barrier (BBB) and subsequent neurological deficits are the most severe consequence of intracerebral hemorrhage (ICH). Minocycline has been wildly used clinically as a neurological protective agent in clinical practice. However, the underlying mechanisms by which minocycline functions remain unclear. Therefore, we assessed the influence of minocycline on BBB structure, neurological function, and inflammatory responses in a collagenase-induced ICH model, and elucidated underlying molecular mechanisms as well. Following a single injection of collagenase VII-S into the basal ganglia, BBB integrity was assessed by Evans blue extravasation while neurological function was assessed using an established neurologic function scoring system. Minocycline treatment significantly alleviated the severity of BBB disruption, brain edema, and neurological deficits in ICH model. Moreover, minocycline decreased the production of inflammatory mediators including TNF, IL-6, and MMP-9, by microglia. Minocycline treatment decreased DKK1 expression but increased Wnt1, β-catenin and Occludin, a phenomenon mimicked by DKK1 silencing. These data suggest that minocycline improves the consequences of ICH by preserving BBB integrity and attenuating neurologic deficits in a DKK1-related manner that involves enhancement of the Wnt1-β-catenin activity., (Copyright © 2019 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: Uterine fibroids are a significant health problem. These common benign tumors occur in 70-80% of women before age 50 years and often cause bleeding and pain and can interfere considerably with daily life. Current treatment options are limited. Fibroids contain substantial amounts of altered and disordered collagens, which contribute to their bulk. Targeting these collagens directly presents a novel treatment approach., Objectives: We sought to test the hypothesis that a highly purified collagenase Clostridium histolyticum will digest interstitial collagen in uterine fibroids and reduce their stiffness and thereby evaluate the feasibility that this collagenase C histolyticum can be developed into an alternative treatment for fibroids. A secondary objective was to describe the collagen content of the fibroid tissue., Study Design: Fibroid tissue cubes (1 cm 3 ; n = 154) were cut from 17 uterine fibroids that were obtained from 7 consented subjects undergoing scheduled hysterectomies. Tissue cubes were injected with diluent, placebo, or highly purified collagenase C histolyticum (0.05, 0.1, or 0.2 mg/cube) and incubated at 37°C for 24, 48, 72, or 96 hours. At each time point, 6 noninjected control cubes were also evaluated. Tissue cubes were photographed before and after incubation. Myometrial samples (n = 21) were also evaluated. Stiffness was quantified through rheometry by measuring complex shear moduli of the tissues. Percent fibrosis was determined by computerized analysis of Masson-trichrome-stained slides. Digestion of collagen fibrils was confirmed by transmission electron microscopy., Results: Fibrosis in untreated fibroids ranged from 37% to 77%, reflecting the collagen-rich nature of these tumors. After treatment with collagenase for 96 hours, fibrosis ranged from 5.3% to 2.4%. Transmission electron microscopy confirmed complete digestion of collagen fibrils. Tissue stiffness was reduced with all 3 doses of collagenase treatment and at all 4 time points. Longer incubation times with collagenase caused greater reduction in stiffness, and treated cubes lost their cuboidal shape and had gelatinous/liquefied centers. At 96 hours the stiffness in tissues treated with the lowest dose was reduced to 966 ± 106 Pascal compared with the diluent-treated control at the same time (5323 ± 903 Pascal; P < .0001; by analysis of variance with Tukey-Kramer)., Conclusion: Uterine fibroids have a high content of collagen that can be effectively digested by highly purified collagenase C histolyticum, resulting in reduced tissue stiffness. Loss of stiffness may decrease bulk symptoms in vivo and possibly lead to shrinkage of fibroids through changed mechanotransduction, leading ultimately to reduced fibroid symptoms of pain and bleeding. Clinical trials are necessary to evaluate the safety and efficacy of collagenase C histolyticum including the rate of regrowth of fibroids. The data of this study provide a strong rationale for using this purified collagenase in clinical trials as a local treatment for women with fibroids., (Copyright © 2016 Elsevier Inc. All rights reserved.)

Background: The fibrotic capsule that surrounds silicone implants consists mainly of collagen. The FDA-approved collagenase of the bacterium clostridium histolyticum provides a reasonable treatment option. Safety and efficacy at the female breast site must be evaluated before clinical utilization., Materials and Methods: We incubated 20 samples of fibrotic capsule as well as 12 full thickness skin grafts harvested from the female breast site for 24 hours with different doses of collagenase. Outcome measures involved histological assessment of thickness and density of the capsule tissue as well as the skin grafts. Furthermore, we performed a collagen assay and immunohistochemistry staining for collagen subtypes., Results: Collagenase treatment was able to degrade human capsule contracture tissue ex-vivo. The remaining collagen subtype after degradation was type 4 only. 0.3 mg/ml of collagenase was most effective in reducing capsule thickness when compared with higher concentrations. Of note, effectiveness was inversely related to capsule density, such that there was less reduction in thickness with higher capsule densities and vice versa. Furthermore, the application of 0.3mg/ml collagenase did not lead to thinning or perforation of full thickness skin grafts., Conclusion: Adjustment of collagenase dose will depend on thickness and density of the contracted capsule. A concentration of 0.3mg/ml seems to be safe and effective in an ex-vivo setting. The remaining collagen subtype 4 is suitable to serve as a neo-capsule/acellular tissue matrix. Collagenase treatment for capsular contracture may soon become a clinical reality.

Objective: To compare the effects of exogenous enzymes on the degradation of adhesive-dentin interface., Methods: Forty molars were sectioned to expose the middle-coronal dentin surface and randomly divided into two adhesive systems: an etch-and-rinse adhesive Adper Single Bond 2 and a self-etching adhesive G-Bond. After composite building up, the specimens were then randomly assigned to four groups(n=5 for each group)as follows: group 1, 24 h of water storage(the control group); group 2, six months of water storage; group 3, twelve weeks storage in artificial saliva containing clostridium histolyticum collagenase; group 4, twelve weeks storage in artificial saliva containing cholesterolesterase. The microtensile bond strengths(MTBS)were then tested. The failure modes and nanoleakage were analyzed., Results: After aging treatments, the three aging groups showed significantly lower MTBS compared with the control group in both adhesive systems(P<0.05). For etch-and-rinse adhesive Adper Single Bond 2, the MTBS of group 3([19.6±3.5]MPa)was lower than that of group 2([23.4±4.2]MPa)and group 4([24.2±4.2]MPa)(P<0.05). For self-etching adhesive G-Bond, there was no difference on MTBS among different aging groups(P>0.05). SEM observation showed that, compared with the control group, water storage(group 2)and the exogenous enzymes(group 3 and 4)increased the nanoleakage expression(silver deposition)of both adhesive systems. Adhesive failure was the predominant fracture modes in all groups., Conclusions: Storage in artificial saliva containing clostridium histolyticum collagenase or cholesterol esterase could be used to accelerate the degradation process of adhesive-dentine interface.

The clostridial collagenases, H and G, play key roles in pancreatic islet isolation. Collagenases digest the peptide bond between Yaa and the subsequent Gly in Gly-Xaa-Yaa repeats. To fully understand the pancreatic islet isolation process, identification of the collagenase substrates in the tissue is very important. Although collagen types I and III were reported as possible substrates for collagenase H, the substrate for collagenase G remains unknown. In this study, collagen type V was focused upon as the target for collagenases. In vitro digestion experiments for collagen type V were performed and analyzed by SDS-PAGE and mass spectrometry. Porcine pancreatic tissues were digested in vitro under three conditions and observed during digestion. The results revealed that collagen type V was only digested by collagenase G and that the digestion was initiated from the N-terminal part. Tissue degradation during porcine islet isolation was only observed in the presence of both collagenases H and G. These findings suggest that collagen type V is one of the substrates for collagenase G. The enzymatic activity of collagenase G appears to be more important for pancreatic islet isolation in large mammals such as pigs and humans.

Multiple organ failure and pancreatic necrosis are the factors that determine prognosis in acute pancreatitis attacks. We investigated the effects of collagenase on the debridement of experimental pancreatic necrosis. The study covered 4 groups; each group had 10 rats. Group I was the necrotizing pancreatitis group. Group II was the collagenase group with pancreatic loge by isotonic irrigation following necrotizing pancreatitis. Group III was the collagenase group with pancreatic loge following necrotizing pancreatitis. Group IV was the intraperitoneal collagenase group following necrotizing pancreatitis. The progress of the groups was compared hematologically and histopathologically. There was no difference among the groups regarding the levels of leukocyte, hemogram, and urea. The differences in AST levels between Group I and II; and differences in glucose, calcium, LDH, AST, and amylase between Group II and III; between Group II and IV; between Group I and III; and between Group I and IV were statistically significant (P < 0.05). There were statistically significant differences between Group II and III, and Group II and IV regarding edema, acinar necrosis, inflammatory cell infiltration, hemorrhage, and fat necrosis (P < 0.05). In conclusion, the collagenase preparation used in this experimental pancreatitis model was found to be effective in the debridement of pancreatic necrosis.

Objective: To investigate the inhibition effect of dopamine on the activity of matrix metalloproteinases (MMP) and the effect of dopamine on degradation of dentin collagen for its potential use in caries treatment and dentin adhesive., Methods: In the experiment of MMP activity test, 2.0 g/L dopamine + 1.0 g/L highly purified collagenase type VIII from Clostridium histolyticum served as the experimental group, and deionized water + 1.0 g/L highly purified collagenase type VIII from Clostridium histolyticum served as the negative control group, and 2% chlorhexidine + 1.0 g/L highly purified collagenase type VIII from Clostridium histolyticum served as the positive control group, and the mixture volume ratio of the two ingredients in every group was 1:9. After 15 minutes, the enzyme activity of each sample was tested by MMP activity colerimetric quantitative detection kits, and the test was repeated 5 times in each group. In the experiment of collagen degradation, the dentin slices were demineralized with 37% phosphoric acid for 1 min. In sequence, 2 dentin slices were used to observe the morphology, and the remaining 30 dentine slices were randomly divided into three groups (n = 10) according to random number table: the negative control ones were stored in 100 µl deionized water and 900 µl collagenase (7 days, 37 °C), the positive control ones were stored in 100 µl chlorhexidine and 900 µl collagenase (7 days, 37 °C) and the experimental specimens were stored in 100 µl dopamine and 900 µl collagenase (7 days, 37 °C). The degraded collagen was investigated by assaying hydroxyproline. The framework of collagen was evaluated with field emission scanning electron microscope (FE-SEM)., Results: The statistical results of completely random design ANOVA showed that the MMP activity and the amount of degraded collagen of the negative control group [(0.089 ± 0.011) µmol · min⁻¹ · mg⁻¹ and (2 837 ± 201) µg/cm²] were significantly higher than those of the positive control group [(0.038 ± 0.006) µmol · min⁻¹ · mg⁻¹ and (1 288 ± 172) µg/cm²] and the experimental group [(0.030 ± 0.009) µmol · min⁻¹ · mg⁻¹ and (1 389 ± 255) µg/cm²] (P < 0.05). SEM observation indicated that the structural integrity of the collagen network on dentin still existed in experiment samples and positive control groups, however, collagen fibrils were destructed and the structural integrity disappeared in the negative control groups., Conclusions: Dopamine may inhibit MMP activity and reduce the amount of degraded collagen.

Context: Osteoarthritis (OA) has become by far the most common joint disorder. A number of studies using OA animal models have explored the effects of agents that can modulate bone metabolism., Objective: In the present study, we investigated the effect of acetylated derivative of plant alkaloid glaucine (ADG) on experimental OA in mice., Materials and Methods: Arthritis was induced by two intraarticular (i.a.) injections of collaganase. Histopathological changes were observed through hematoxylin and eosine (H&E), safranin O and toluidine blue staining. Differentiation of bone marrow (BM) cells was evaluated by tartarate-resistant acid phosphatase (TRAP) assay. The expression of phospho-Janus kinase 2 (pJAK2) and phospho signal transducer and activator of transcription3 (pSTAT3) expression in the joints was determined by immunohistochemistry., Results: We established that ADG significantly decreased cell infiltration (2.32 ± 0.14 versus 1.62 ± 0.13), cartilage loss (2.42 ± 0.12 versus 1.12 ± 0.10) and bone erosion (1.76 ± 0.13 versus 1.04 ± 0.14) in arthritic mice. It appeared that the substance inhibited in a dose-dependent manner osteoclast differentiation in vitro. ADG suppressed the expression of pJAK2 in the joint and partially affected the expression of pSTAT3., Conclusion: Present results suggest that ADG is a suitable candidate for further development as an anti-arthritic agent.