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Identification of myeloperoxidase, alpha-defensin and calgranulin in calcium oxalate renal stones.
Clin Chim Acta. 2007 Sep; 384(1-2):41-7.CC

Abstract

BACKGROUND

In order to understand the mechanism of stone genesis, it is essential to determine the characteristics of macromolecules constituting the urinary stones. We characterized proteins from the inner core and outer matrix of calcium oxalate (CaOx) renal stones.

METHODS

Inner core and outer matrix of CaOx renal stones were separated and proteins were extracted with a buffer containing SDS and beta-mercaptoethanol. Proteins were analyzed and purified by SDS-PAGE and RP-HPLC respectively. The protein bands from gel and protein fractions were sequenced by MALDI TOF mass spectrometry. ELISA, western and slot blot immunoassays were performed to confirm the identity of the proteins in stones and urine of the stone formers. The potential of the identified protein as an effective promoter or inhibitor was assessed by observing their effects on CaOx crystallization using aggregometer.

RESULTS

The inner core extract predominantly exhibited protein species in the molecular weight range of 12-14 kDa. However, a 66 kDa band, identified as osteopontin was also detected in the inner core along with outer matrix and in the urine of stone formers and non stone formers. Purification of low molecular weight proteins was carried out by reversed phase HPLC. Tandem mass spectrometry analysis identified them as myeloperoxidase chain A (MPO-A), alpha-defensin, and calgranulin. ELISA, western blot and slot-blot immuno-assays further confirmed their presence restricted to the inner core and not in the outer matrix. Turbidity assays showed that low molecular weight renal stone proteins promoted the aggregation of CaOx crystals.

CONCLUSIONS

Persistent hyperoxaluria leads to tubular epithelial injury, resulting in the release of these anti-inflammatory proteins. These proteins could have been first adsorbed on CaOx crystals thereby become a part of nucleation process leading to inner matrix formation.

Authors+Show Affiliations

Department of Biological and Biomedical Sciences, Research Laboratory, Juma Building, Pakistan.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

17610860

Citation

Mushtaq, Shamim, et al. "Identification of Myeloperoxidase, Alpha-defensin and Calgranulin in Calcium Oxalate Renal Stones." Clinica Chimica Acta; International Journal of Clinical Chemistry, vol. 384, no. 1-2, 2007, pp. 41-7.
Mushtaq S, Siddiqui AA, Naqvi ZA, et al. Identification of myeloperoxidase, alpha-defensin and calgranulin in calcium oxalate renal stones. Clin Chim Acta. 2007;384(1-2):41-7.
Mushtaq, S., Siddiqui, A. A., Naqvi, Z. A., Rattani, A., Talati, J., Palmberg, C., & Shafqat, J. (2007). Identification of myeloperoxidase, alpha-defensin and calgranulin in calcium oxalate renal stones. Clinica Chimica Acta; International Journal of Clinical Chemistry, 384(1-2), 41-7.
Mushtaq S, et al. Identification of Myeloperoxidase, Alpha-defensin and Calgranulin in Calcium Oxalate Renal Stones. Clin Chim Acta. 2007;384(1-2):41-7. PubMed PMID: 17610860.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Identification of myeloperoxidase, alpha-defensin and calgranulin in calcium oxalate renal stones. AU - Mushtaq,Shamim, AU - Siddiqui,Anwar Ali, AU - Naqvi,Zulfiqar Ali, AU - Rattani,Ahmed, AU - Talati,Jamsheer, AU - Palmberg,Carina, AU - Shafqat,Jawed, Y1 - 2007/05/26/ PY - 2006/09/29/received PY - 2007/03/19/revised PY - 2007/05/15/accepted PY - 2007/7/6/pubmed PY - 2007/12/6/medline PY - 2007/7/6/entrez SP - 41 EP - 7 JF - Clinica chimica acta; international journal of clinical chemistry JO - Clin Chim Acta VL - 384 IS - 1-2 N2 - BACKGROUND: In order to understand the mechanism of stone genesis, it is essential to determine the characteristics of macromolecules constituting the urinary stones. We characterized proteins from the inner core and outer matrix of calcium oxalate (CaOx) renal stones. METHODS: Inner core and outer matrix of CaOx renal stones were separated and proteins were extracted with a buffer containing SDS and beta-mercaptoethanol. Proteins were analyzed and purified by SDS-PAGE and RP-HPLC respectively. The protein bands from gel and protein fractions were sequenced by MALDI TOF mass spectrometry. ELISA, western and slot blot immunoassays were performed to confirm the identity of the proteins in stones and urine of the stone formers. The potential of the identified protein as an effective promoter or inhibitor was assessed by observing their effects on CaOx crystallization using aggregometer. RESULTS: The inner core extract predominantly exhibited protein species in the molecular weight range of 12-14 kDa. However, a 66 kDa band, identified as osteopontin was also detected in the inner core along with outer matrix and in the urine of stone formers and non stone formers. Purification of low molecular weight proteins was carried out by reversed phase HPLC. Tandem mass spectrometry analysis identified them as myeloperoxidase chain A (MPO-A), alpha-defensin, and calgranulin. ELISA, western blot and slot-blot immuno-assays further confirmed their presence restricted to the inner core and not in the outer matrix. Turbidity assays showed that low molecular weight renal stone proteins promoted the aggregation of CaOx crystals. CONCLUSIONS: Persistent hyperoxaluria leads to tubular epithelial injury, resulting in the release of these anti-inflammatory proteins. These proteins could have been first adsorbed on CaOx crystals thereby become a part of nucleation process leading to inner matrix formation. SN - 0009-8981 UR - https://www.unboundmedicine.com/medline/citation/17610860/Identification_of_myeloperoxidase_alpha_defensin_and_calgranulin_in_calcium_oxalate_renal_stones_ L2 - https://linkinghub.elsevier.com/retrieve/pii/S0009-8981(07)00292-6 DB - PRIME DP - Unbound Medicine ER -