Free radical generation during chemical depolymerization of heparin

Rota, Cristina; Liverani, Lino; Spelta, Franco; Mascellani, Giuseppe; Tomasi, Aldo; Iannone, Anna; Vismara, Elena

doi:10.1016/j.ab.2005.06.043

Cited by 24 publications

(18 citation statements)

References 20 publications

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“…In vitro, exposure of rat glomerular BM HS to ROS generated by hypoxanthine/xanthine oxidase reactions results in HS fragmentation or depolymerization, detected by PAGE (34). Similarly, hydroxyl radicals have previously been shown to efficiently depolymerize heparin to low-molecular weight heparins (35). Alternatively, free radicals may be chemically detoxified by removing the anomeric hydrogen of internal sugar residues of HS, as previously reported for polyelectrolyte configurations of glucose, such as phosphorylated or sulfated glucans (36).…”

Section: Discussionmentioning

confidence: 76%

“…Overall, these findings suggest that in situ, normal levels of endogenous intracellular HS may function, at least in part, to protect β cells from free radical-induced cell injury. Heparin imported into β cells in vitro and endogenous β cell HS in situ (i.e., in the pancreas) may interact directly with ROS and function as an antioxidant or free radical sink (34)(35)(36). The depolymerization of HS by free radical species, for example, could represent an antioxidant mechanism for protecting β cells from free radical damage.…”

Section: Discussionmentioning

confidence: 99%

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Heparan sulfate and heparanase play key roles in mouse β cell survival and autoimmune diabetes

Ziolkowski

Popp²,

Freeman³

et al. 2012

J. Clin. Invest.

145

200

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The autoimmune type 1 diabetes (T1D) that arises spontaneously in NOD mice is considered to be a model of T1D in humans. It is characterized by the invasion of pancreatic islets by mononuclear cells (MNCs), which ultimately leads to destruction of insulin-producing β cells. Although T cell dependent, the molecular mechanisms triggering β cell death have not been fully elucidated. Here, we report that a glycosaminoglycan, heparan sulfate (HS), is expressed at extraordinarily high levels within mouse islets and is essential for β cell survival. In vitro, β cells rapidly lost their HS and died. β Cell death was prevented by HS replacement, a treatment that also rendered the β cells resistant to damage from ROS. In vivo, autoimmune destruction of islets in NOD mice was associated with production of catalytically active heparanase, an HS-degrading enzyme, by islet-infiltrating MNCs and loss of islet HS. Furthermore, in vivo treatment with the heparanase inhibitor PI-88 preserved intraislet HS and protected NOD mice from T1D. Our results identified HS as a critical molecular requirement for islet β cell survival and HS degradation as a mechanism for β cell destruction. Our findings suggest that preservation of islet HS could be a therapeutic strategy for preventing T1D.

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Section: Discussionmentioning

confidence: 76%

Section: Discussionmentioning

confidence: 99%

Heparan sulfate and heparanase play key roles in mouse β cell survival and autoimmune diabetes

Ziolkowski

Popp²,

Freeman³

et al. 2012

J. Clin. Invest.

145

200

View full text Add to dashboard Cite

show abstract

“…detecting the O-sulfate and N-sulfate groups) and hence proportional to the negative charge carried by the HS chains. In contrast, the lack of Alcian blue staining of isolated C57BL/6 islets (C) indicates the loss of highly sulfated HS, consistent with oxidant-mediated HS chain depolymerization (see Figure 9, Figure S3) (52,53). For immunohistochemistry, antigen retrieval in paraffin sections was achieved using 0.05% pronase and HS was localized using 10E4 mouse-anti-human HS mAb (Amsbio, UK), horseradish peroxidase-conjugated rabbit-anti-mouse Ig and AEC (as the chromogen).…”

Section: Supporting Informationmentioning

confidence: 86%

“…Reactive chemical species have been reported to depolymerize HS and HS analogs such as heparin (52,53). To evaluate the contribution of oxidative damage to the loss of islet HS during islet isolation, we pretreated the donor mice with the antioxidant BHA (120 mg/kg i.p.)…”

Section: Antioxidant Treatment Preserves Islet Hs During Islet Isolationmentioning

confidence: 99%

Islet Heparan Sulfate but Not Heparan Sulfate Proteoglycan Core Protein Is Lost During Islet Isolation and Undergoes Recovery Post-Islet Transplantation

Choong

Freeman

Parish

et al. 2015

American Journal of Transplantation

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Islet beta cells in situ express intracellular heparan sulfate (HS), a property previously shown in vitro to be important for their survival. We report that HS levels inside islet beta cells correlate with the novel intracel-lular localization of the HSPG core proteins for collagen type XVIII (Col18), a conventional extracellular matrix component. Syndecan-1 (Sdc1) and CD44 core proteins were similarly localized inside beta cells. During isolation, mouse islets selectively lose HS to 11-27% of normal levels but retain their HSPG core proteins. Intra-islet HS failed to recover substantially during culture for 4 days and was not reconstituted in vitro using HS mimetics. In contrast, significant recovery of intra-islet HS to $40-50% of normal levels occurred by 5-10 days after isotransplantation. Loss of islet HS during the isolation procedure is independent of heparanase (a HS-degrading endoglycosidase) and due, in part, to oxidative damage. Treatment with antioxidants reduced islet cell death by $60% and increased the HS content of isolated islets by $twofold compared to untreated islets, preserving intra-islet HS to $60% of the normal HS content of islets in situ. These findings suggest that the preservation of islet HS during the islet isolation process may optimize islet survival posttransplant. Abbreviations: AEC, 3-amino-9-ethylcarbazole; A.U., arbitrary units; B6.Hpse-KO, heparanase knockout C57BL/6; BHA, butylated hydroxyanisole; BM, basement membrane; Col18, collagen type XVIII; DMTU, dimethylthiourea; ECM, extracellular matrix; EXT1-2, exostosin 1-2; EXTL1-3, exostosin-like 1-3; FITC, fluo-rescein isothiocyanate; GMFR, geometric mean fluo-rescence ratio; Gpc2, glypican-2; H&E, haematoxylin and eosin; HS, heparan sulfate; HSPG, heparan sulfate proteoglycan; MFI, mean fluorescence intensity; MNCs, mononuclear cells; Ndst2, N-deacetylase/N-sulfotransferase 2; NOD/Lt, nonobese diabetic; PSN, penicillin-streptomycin-neomycin; qRT-PCR, quantitative reverse transcription polymerase chain reaction; ROS, reactive oxygen species; Sdc1, syndecan-1; SEM, standard error of the mean

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“…While this double bond can be eliminated from heparosan by deleting the gene of K5 lyase from the K5 Escherichia coli genome, this would result in a decrease of heparosan in the culture supernatant and an increase in heparosan chain size. While it is possible to control chain size and remove the unsaturated sugar from heparosan by chemical methods using hydrogen peroxide or ozone, 36 metabolic engineering offers an alternative solution. The double bond can also be removed with Δ4,5-glycuronidase, an unusual enzyme that hydrolyzes the unsaturated Δ4,5 uronic acid at the nonreducing end of polysaccharides.…”

mentioning

confidence: 99%

Escherichia coliK5 heparosan fermentation and improvement by genetic engineering

Wang¹,

Dordick²,

Linhardt³

2011

Bioengineered Bugs

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Free radical generation during chemical depolymerization of heparin

Cited by 24 publications

References 20 publications

Heparan sulfate and heparanase play key roles in mouse β cell survival and autoimmune diabetes

Heparan sulfate and heparanase play key roles in mouse β cell survival and autoimmune diabetes

Islet Heparan Sulfate but Not Heparan Sulfate Proteoglycan Core Protein Is Lost During Islet Isolation and Undergoes Recovery Post-Islet Transplantation

Escherichia coliK5 heparosan fermentation and improvement by genetic engineering

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