The Effect of Truncated Collagenase Class I Isomers on Human Islet Isolation Outcome

Brandhorst, Heide; Asif, Sana; Andersson, Karin; Mönch, Johanna; Friedrich, O; Rämsch-Günther, Nicole; Rämsch, Christian; Steffens, Melanie C.; Lambrecht, J. Thomas; Schräder, Thomas; Kurfürst, Manfred; Andersson, Helene H.; Felldin, Marie; Foss, Aksel; Salmela, K.; Tibell, Annika; Tufveson, Gunnar; Korsgren, Olle; Brandhorst, Daniel

doi:10.1097/tp.0b013e3181e49bd7

Cited by 12 publications

(9 citation statements)

References 8 publications

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“…Previous studies in the rat identified CC-II as the most relevant isoform for pancreas digestion when compared with CC-I [47,59]. In agreement with the observations of the Groningen group [60], we found that human pancreases are efficiently digested even in the absence of CC-I as long as sufficient activities of supplementary neutral proteases are present [61,62]. Nevertheless, for optimal collagenase function, a certain amount of CC-I has to be present within the enzyme blend.…”

Section: Enzyme Characterizationsupporting

confidence: 90%

“…The loss of intact CC-I does not only have an effect on the CC-II-to-CC-I ratio as an important variable for collagenase efficiency [63,64], it also determines the amount of neutral proteases required for efficient pancreas dissociation [67]. In agreement with observations by the Groningen group [60], we found that human pancreases are efficiently digested, even in the absence of intact CC-I, as long as sufficient activities of supplementary proteases such as CNP or clostripain are present [61,62]. While CNP or TL may compensate the reduced activity of CC-I, these enzymes have a detrimental effect on islet viability, as discussed in section 3.…”

Section: Collagenase Degradationsupporting

confidence: 88%

See 1 more Smart Citation

Enzyme Development for Human Islet Isolation: Five Decades of Progress or Stagnation?

Brandhorst¹,

Brandhorst²,

Prv.³

2017

Rev Diabet Stud

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In comparison to procedures used for the separation of individual cell types from other organs, the process of human pancreatic islet isolation aims to digest the pancreatic exocrine matrix completely without dispersing the individual cells within the endocrine cell cluster. This objective is unique within the field of tissue separation, and outlines the challenge of islet isolation to balance two opposing priorities. Although significant progress has been made in the characterization and production of enzyme blends for islet isolation, there are still numerous areas which require improvement. The ultimate goal of enzyme production, namely the routine production of a consistent and standardized enzyme blend, has still not been realized. This seems to be mainly the result of a lack of detailed knowledge regarding the structure of the pancreatic extracellular matrix and the synergistic interplay between collagenase and different supplementary proteases during the degradation of the extracellular matrix. Furthermore, the activation of intrinsic proteolytic enzymes produced by the pancreatic acinar cells, also impacts on the chance of a successful outcome of human islet isolation. This overview discusses the challenges of pancreatic enzymatic digestion during human islet isolation, and outlines the developments in this field over the past 5 decades.

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Section: Enzyme Characterizationsupporting

confidence: 90%

Section: Collagenase Degradationsupporting

confidence: 88%

Enzyme Development for Human Islet Isolation: Five Decades of Progress or Stagnation?

Brandhorst¹,

Brandhorst²,

Prv.³

2017

Rev Diabet Stud

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“…The aim was to prove our hypothesis that NPs can compensate reduced CI activity to isolate significant amounts of islets from the human pancreas. To be consistent with our previous study about the effect of intact and degraded CI on islet isolation outcome, 17 NP and CP were combined to serve as complementary proteases.…”

mentioning

confidence: 71%

Comparison of Neutral Proteases and Collagenase Class I as Essential Enzymes for Human Islet Isolation

Brandhorst

Kurfürst²,

Johnson

et al. 2016

Transplantation Direct

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BackgroundEfficient islet isolation requires synergistic interaction between collagenase class I (CI) and class II (CII). The CI degradation alters the ratio between CI and CII and is responsible for batch-to-batch variations. This study compares the role of neutral protease (NP) plus clostripain (CP) with CI as essential enzymes for human islet isolation.MethodsHuman islets were isolated using 4 different enzyme mixtures composed of CII plus either intact (CI-115) or degraded CI (CI-100). Blends were administered either with or without NP/CP. Purified islets were cultured for 3 to 4 days before islet quality assessment.ResultsWhereas using intact CI-115 without NP/CP did not significantly reduce islet yield (3429 ± 631 vs 3087 ± 970 islet equivalent/g, nonsignificant), administration of degraded CI-100 without NP/CP decreased islet yield from 3501 ± 580 to 1312 ± 244 islet equivalent/g (P < 0.01), doubled the amount of undigested tissue from 11.8 ± 1.6 to 24.4 ± 1.2% (P < 0.01) and triplicated the percentage of trapped islets from 7.7 ± 2.8 to 22.5 ± 3.6% (P < 0.05). Islet yield did not vary between supplemented CI-115 and CI-100, but was increased using CI-115 when NP/CP was omitted (P < 0.05). A trend toward higher viability and increased secretory insulin response was noted in both CI-100 and CI-115 when NP/CP was not added.ConclusionsThis study suggests that NP/CP can compensate reduced CI activity. Future attempts to optimize enzyme blends should consider the possibility to increase the proportion of collagenase CI to reduce the need for potentially harmful NPs.

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“…For instance, the enzymes of C. histolyticum were recently approved as drugs to break down the tough cords in morbus Dupuytren (15–16) and are widely used in human cell islet isolation (17, 18) and wound debridement (19). …”

Section: Introductionmentioning

confidence: 99%

Structural Basis for Activity Regulation and Substrate Preference of Clostridial Collagenases G, H, and T

Eckhard

Schönauer

Brandstetter

2013

Journal of Biological Chemistry

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Background: Bacterial collagenases degrade collagen substrates with high efficiency yet varying specificity.Results: The newly identified calcium site, aspartate switch, and conformational selectivity filter regulate substrate access to the active sites of these collagenases.Conclusion: The unanticipated dynamics of the substrate recognition sites plus zinc occupancy combine to tune the enzymatic activity.Significance: The crystal structures provide a rational framework to understand and optimize the isoform-dependent collagenase activities.

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The Effect of Truncated Collagenase Class I Isomers on Human Islet Isolation Outcome

Cited by 12 publications

References 8 publications

Enzyme Development for Human Islet Isolation: Five Decades of Progress or Stagnation?

Enzyme Development for Human Islet Isolation: Five Decades of Progress or Stagnation?

Comparison of Neutral Proteases and Collagenase Class I as Essential Enzymes for Human Islet Isolation

Structural Basis for Activity Regulation and Substrate Preference of Clostridial Collagenases G, H, and T

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