Histochemical Analysis of the Role of Class I and Class II Clostridium Histolyticum Collagenase in the Degradation of Rat Pancreatic Extracellular Matrix for Islet Isolation

Vos-Scheperkeuter, Greetje H.; Suylichem, P.T.R. van; Vonk, Maike W.A.; Wolters, G. H. J.; Schilfgaarde, R. van

doi:10.1177/096368979700600407

Cited by 23 publications

(18 citation statements)

References 18 publications

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“…Study by Brandhorst et al [12] showed that neither C1 nor C2 alone were able to dissociate pancreatic tissue. Rapid and complete loss of peri-islet ECM depends on both collagenase isoform and is best achieved with assistance of other proteolytic enzymes [10] [12] [13]. This conclusion was also confirmed in our study, where the best results were achieved with a batch from Group 3 containing intact and fragmented collagenase in combination with neutral protease (NP) and clostripain.…”

Section: Discussionsupporting

confidence: 81%

“…Clostridium collagenases C1 and C2 hydrolyze collagen chains sequentially from the ends into mixture of small peptides [9]. It was reported that C2 was more efficient in collagen degradation [10] whereas C1 played only supporting role in islet isolations [11]. Study by Brandhorst et al [12] showed that neither C1 nor C2 alone were able to dissociate pancreatic tissue.…”

Section: Discussionmentioning

confidence: 99%

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Testing of a New Collagenase Blend for Pancreatic Islet Isolation Produced by <i>Clostridium histolyticum</i>

Berková¹,

Saudek²,

Leontovyč³

et al. 2018

ABB

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Clostridium histolyticum is used for production of several proteolytic enzymes such as elastase, neutral proteases, clostripain and in particular collagenase. Besides industrial applications, collagenase has been indispensable for medical purposes including isolation of pancreatic islets for diabetes treatment. The aim of this study was to optimize the method for production and partial purification of a new collagenase blend and to test its suitability for successful pancreatic islets isolation in a rat model. Bacterial strain of C. histolyticum was sequenced for presence of the collagenase genes. Different fermentation conditions were tested and the process of collagenase extraction was modified and optimized. Samples of collagenases were taken for western blot detection, activity assessment, and ability for dissociation of pancreatic tissue. Findings indicate that concentrated trypton growth medium with pepton was the most suitable for Clostridium growth and collagenase production.Whole genome sequencing revealed two genes for collagenase and also gene for clostripain. Western blot specific detection helped to select useful production modifications. Following these modifications was also improved the yield, morphology and in vitro function of intact pancreatic islets which were finally comparable or better than those achieved using standard blends of collagenase. The results support the use of the new collagenase blend for islet isolation giving thus the opportunity to choose an alternative product. Our next steps would lead to further enzyme purification through scaling up of the production method for a wider use.

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Section: Discussionsupporting

confidence: 81%

Section: Discussionmentioning

confidence: 99%

Testing of a New Collagenase Blend for Pancreatic Islet Isolation Produced by <i>Clostridium histolyticum</i>

Berková¹,

Saudek²,

Leontovyč³

et al. 2018

ABB

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“…Concerning the rat pancreas digestion, it has been reported that the CII is the most efficient at digesting exocrine tissue, whereas CI increases the digestion rate (13,14); therefore, an appropriate mix of CI and CII has been proposed to optimize islet isolation (15)(16)(17)(18). In the early stages of digestion, neutral proteases also are needed, but in excessive amounts these can destroy islet integrity (19 -21).…”

mentioning

confidence: 99%

Characterization of Collagenase Blend Enzymes for Human Islet Transplantation

Antonioli¹,

Fermo

Cainarca

et al. 2007

Transplantation

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Background. Efficient islet isolation represents a necessary requirement for successful islet transplantation as a treatment for type 1 diabetes. The choice of collagenase for pancreas digestion is critical for the isolation outcome, and Liberase™ is the most widely used enzyme, although large intra-batched variability in activity and efficiency has been observed. Methods. The aim of this study was to characterize Liberase™ components and their relative role in pancreas digestion. Liberase batches were characterized by microelectrophoresis. Results. By means of microelectrophoresis, we identified three main proteins each with different prevalences between batches. Two proteins were found to correspond to class I (CI) and one to class II (CII) collagenase. In a series of 163 islet isolations, we observed that the CII correlated with islet yield (PϽ0.001) and digestion time (PϽ0.001); additionally, CI directly correlated with purity (Pϭ0.028). Finally, when CII and one of the CI isoforms were Ͼ50 percentile, 15 of 36 preparations were transplanted, with 27 of 127 transplanted in the other cases (Pϭ0.013). Conclusion. These results represent an important step toward the characterization of enzymes, with the final aim of identifying key components for a standardized product.

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“…Collagenase II was shown to be important for rat pancreatic islet isolation, 6,12 and a major role for collagenase I has been demonstrated for human islet isolation. 8 More importantly, in the latter study, collagenase Ib was shown to reduce islet yields, and a correlation between degraded collagenase and reduced islet viability has been previously demonstrated.…”

Section: Discussionmentioning

confidence: 99%

A new collagenase blend increases the number of islets isolated from mouse pancreas

Yeşil¹,

Michel²,

Chwalek³

et al. 2009

Islets

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Histochemical Analysis of the Role of Class I and Class II Clostridium Histolyticum Collagenase in the Degradation of Rat Pancreatic Extracellular Matrix for Islet Isolation

Abstract: Histochemical analysis of the role of class I and class II Clostridium histolyticum collagenase in the degradation of rat pancreatic extracellular matrix for islet isolation.

Cited by 23 publications

References 18 publications

Testing of a New Collagenase Blend for Pancreatic Islet Isolation Produced by <i>Clostridium histolyticum</i>

Testing of a New Collagenase Blend for Pancreatic Islet Isolation Produced by <i>Clostridium histolyticum</i>

Characterization of Collagenase Blend Enzymes for Human Islet Transplantation

A new collagenase blend increases the number of islets isolated from mouse pancreas

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Histochemical Analysis of the Role of Class I and Class II Clostridium Histolyticum Collagenase in the Degradation of Rat Pancreatic Extracellular Matrix for Islet Isolation

Abstract: Histochemical analysis of the role of class I and class II Clostridium histolyticum collagenase in the degradation of rat pancreatic extracellular matrix for islet isolation.

Cited by 23 publications

References 18 publications

Testing of a New Collagenase Blend for Pancreatic Islet Isolation Produced by &lt;i&gt;Clostridium histolyticum&lt;/i&gt;

Testing of a New Collagenase Blend for Pancreatic Islet Isolation Produced by &lt;i&gt;Clostridium histolyticum&lt;/i&gt;

Characterization of Collagenase Blend Enzymes for Human Islet Transplantation

A new collagenase blend increases the number of islets isolated from mouse pancreas

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Product

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Testing of a New Collagenase Blend for Pancreatic Islet Isolation Produced by <i>Clostridium histolyticum</i>

Testing of a New Collagenase Blend for Pancreatic Islet Isolation Produced by <i>Clostridium histolyticum</i>