Journal: Calcified Tissue International

Article Title: Bone from Healthy Individuals and Patients with CKD Expresses the Sodium-Glucose Co-transporter-2 (SGLT2)

doi: 10.1007/s00223-026-01498-7

Figure Lengend Snippet: Preliminary qPCR amplification for primer selection. A and B Unsuccessful preliminary qPCR amplification using 267 and 430 bp primer pairs for SLC5A2 for MG-63 cells, respectively. The amplification plot displays the performance designed to amplify as a product of the human SLC5A2 transcript ( NM_003041.4 ), with its specific binding sites shown below. While the primers yielded a clear amplification signal in the positive control HK-2 renal cells, no detectable expression was observed in the osteoblast-like MG-63 cell line. Due to the lack of amplification in MG-63 cells, these primer sets were deemed unsuitable for the study. ( C ) Successful validation of the selected 108 bp primer for SLC5A2. This primer pair shows robust and comparable amplification in both MG-63 and HK-2 cell lines between 22 and 24 cycles, confirming its suitability for subsequent comparative gene expression analysis by qPCR

Article Snippet: Human osteoblast-like (MG-63) and human kidney (HK-2) cell lines were obtained from the American Type Culture Collection (ATCC CRL-1427 and -2190, USA).

Techniques: Amplification, Selection, Binding Assay, Positive Control, Expressing, Biomarker Discovery, Gene Expression

Journal: Calcified Tissue International

Article Title: Bone from Healthy Individuals and Patients with CKD Expresses the Sodium-Glucose Co-transporter-2 (SGLT2)

doi: 10.1007/s00223-026-01498-7

Figure Lengend Snippet: Relative quantification of SLC5A2 gene expression. A Raw Ct values of two biologicals (in duplicates) showing comparable SLC5A2 expression in MG-63 and HK-2 cells; for the first time, the SLC5A2 expression in osteoblast-like MG-63 cells is confirmed; B Raw Ct values of three healthy individuals bone samples and ten bone samples from patients with CKD (in duplicates) indicating that the SLC5A2 expression is lower in bone samples from patients with CKD. After ΔΔCt normalization by GAPDH, no significant difference is seen between C cell lines (two biologicals in duplicates), or between D bone samples (mean of duplicates) from healthy subjects and patients with CKD. GAPDH variation in expression (above 3.5 Cts) prevented a suitable normalization of SLC5A2 expression in healthy subjects versus patients with CKD; thus, each condition was normalized to its own GAPDH expression for representation purposes

Article Snippet: Human osteoblast-like (MG-63) and human kidney (HK-2) cell lines were obtained from the American Type Culture Collection (ATCC CRL-1427 and -2190, USA).

Techniques: Quantitative Proteomics, Gene Expression, Expressing

Journal: Calcified Tissue International

Article Title: Bone from Healthy Individuals and Patients with CKD Expresses the Sodium-Glucose Co-transporter-2 (SGLT2)

doi: 10.1007/s00223-026-01498-7

Figure Lengend Snippet: Absolute quantification of SLC5A2 and GAPDH transcript copy numbers in cells (two biologicals in duplicates) and human bone (mean of duplicates from three healthy individuals’ bone and ten samples from patients with CKD) after 40 cycles. The use of standard curves and linear regressions using dilutions in log 10 -3, -4, -5, and -6 from experimental controls (HK-2 cells and apparently healthy individuals´ bone samples) templates allowed the estimation of an unknown number of copies for each primer individually. This analysis confirms comparable absolute SLC5A2 transcript levels between HK-2 and osteoblast-like MG-63 cells, while GAPDH levels were modestly lower in MG-63 cells. In human bone tissue, a non-significant trend towards higher SLC5A2 copies was observed in samples from patients with chronic kidney disease (CKD) compared to samples from healthy subjects. Critically, GAPDH copy numbers were significantly reduced in CKD bone, demonstrating its instability as a reference gene in this pathology and reinforcing the value of absolute quantification for accurate expression analysis while analyzing bone samples. Additionally, internal controls also using pools from biologicals of HK-2 cells and healthy bone (red dashed line indicated within each ‘y’ axis) were adopted as an extra sample (in duplicates) to corroborate the unknown individual sample results; this reference obtained its number of copies within the range of their tested biologicals

Article Snippet: Human osteoblast-like (MG-63) and human kidney (HK-2) cell lines were obtained from the American Type Culture Collection (ATCC CRL-1427 and -2190, USA).

Techniques: Quantitative Proteomics, Expressing