Journal: Cells

Article Title: Canagliflozin, an Inhibitor of the Na + -Coupled D-Glucose Cotransporter, SGLT2, Inhibits Astrocyte Swelling and Brain Swelling in Cerebral Ischemia.

doi: 10.3390/cells12182221

Figure Lengend Snippet: Figure 2. SGLT2 is upregulated in astrocyte cell bodies and astrocyte endfeet following cerebral ischemia. (a–e) Double immunolabeling for SGLT2 (green) and NeuN (a,b) or GFAP (c–e) (red) in contralateral (Contra) and ipsilateral (Ipsi) brain sections from mice post-MCAo/R (2/6 h); merged images are shown in a, b, d, e; individual labelings are shown in c, d—inset, e—inset; the images shown are representative of findings in 3 mice; scale bars, 25 µm except insert, 10 µm.

Article Snippet: We studied SGLT2 expression using a commercial rabbit polyclonal anti-SGLT2 antibody (#NBP1-92384; Novus Biologicals, Centennial, CO, USA).

Techniques: Immunolabeling

Journal: Cells

Article Title: Canagliflozin, an Inhibitor of the Na + -Coupled D-Glucose Cotransporter, SGLT2, Inhibits Astrocyte Swelling and Brain Swelling in Cerebral Ischemia.

doi: 10.3390/cells12182221

Figure Lengend Snippet: Figure 3. Slc5a2 mRNA and SGLT2 protein are upregulated in astrocytes following cerebral ischemia. (a–c) Images (a) and quantification (b,c) of RNAscope for Slc5a2 (red) and Aqp4 (green) mRNA in contralateral (Contra) and ipsilateral (Ipsi) brain sections from mice post-MCAo/R (2/6 h); scale bar, 25 µm; data from 3 mice per group; **, p < 0.01. (d) qPCR for Slc5a2 mRNA in astrocytes and neurons isolated from the contralateral (Contra) and ipsilateral (Ipsi) MCA territory of mice following MCAo/R (2/6 h) and uninjured controls (naïve), expressed as fold-change; 5 mice per group; *, p < 0.05. (e) Quantitative immunohistochemistry for SGLT2 expressed within regions of interest determined by NeuN; data from 7 mice per group; ns, not significant. (f,g) Immunoblot (f) and quantification of immunoblot (g) for SGLT2 in astrocytes isolated from the contralateral (Contra) and ipsilateral (Ipsi) MCA territory of mice following MCAo/R (2/6 h) and uninjured controls (naïve), expressed as fold-change; PC, positive control (kidney lysate); β-actin used as a loading control; 3 mice per group.

Article Snippet: We studied SGLT2 expression using a commercial rabbit polyclonal anti-SGLT2 antibody (#NBP1-92384; Novus Biologicals, Centennial, CO, USA).

Techniques: RNAscope, Isolation, Immunohistochemistry, Western Blot, Positive Control, Control

Journal: Cells

Article Title: Canagliflozin, an Inhibitor of the Na + -Coupled D-Glucose Cotransporter, SGLT2, Inhibits Astrocyte Swelling and Brain Swelling in Cerebral Ischemia.

doi: 10.3390/cells12182221

Figure Lengend Snippet: Figure 4. D-glucose-induced Na+ influx in post-ischemic astrocytes is inhibited by canagliflozin and by the astrocyte-specific deletion of Slc5a2/SGLT2. (a) Live cell image of a tdTomato-expressing astrocyte in an ex vivo brain slice from an MCAo/R (2/6 h) mouse, following incubation with ING-2; scale bar, 25 µm. (b–e) Time course of Na+ influx with step change in D-glucose from 2 to 10 mM in ipsilateral (Ipsi) vs. contralateral (Contra) astrocytes (b); in ipsilateral astrocytes without and with pre-incubation with canagliflozin (5 µM) (c); in ipsilateral astrocytes without and with the addition of canagliflozin (5 µM) at the time indicated by the bar (d); in ipsilateral astrocytes from Ast-Slc5a2WT

Article Snippet: We studied SGLT2 expression using a commercial rabbit polyclonal anti-SGLT2 antibody (#NBP1-92384; Novus Biologicals, Centennial, CO, USA).

Techniques: Expressing, Ex Vivo, Slice Preparation, Incubation

Journal: PeerJ

Article Title: Adrenal SGLT1 or SGLT2 as predictors of atherosclerosis under chronic stress based on a computer algorithm

doi: 10.7717/peerj.15647

Figure Lengend Snippet: (A–B) Rose-colored SGLT1 fluorescence and pink-colored SGLT2 fluorescence could be seen in the adrenal cross sections of the four groups. (C) Semi-quantitative analysis of SGLT1 and SGLT2 expression in the adrenal glands of four groups showed that SGLT1 and SGLT2 expression levels were higher in mice after stress than in the corresponding non-stressed mice. (D) Expression of SGLT1 and SGLT2 was determined by Western blot analysis. (E) mRNA expression of SGLT1 and SGLT2 was detected by qPCR.

Article Snippet: Transverse sections of adrenal glands were incubated with SGLT1 Polyclonal Antibody (1:200, bs-1128R; BIOSS, Beijing, China) and SGLT2 Polyclonal Antibody (1:200, 24654-1-AP; Proteintech, Rosemont, IL, USA).

Techniques: Fluorescence, Expressing, Western Blot

Journal: PeerJ

Article Title: Adrenal SGLT1 or SGLT2 as predictors of atherosclerosis under chronic stress based on a computer algorithm

doi: 10.7717/peerj.15647

Figure Lengend Snippet: CYP11B2 labeled in red showed the adrenal cortex region, while tyrosine hydroxylase (TH) labeled in green showed the adrenal medulla. The nucleus was stained blue by DAPI. SGLT1 was detected by a rose and SGLT2 by a pink. The distribution of SGLT1 and SGLT2 in the adrenal glands of mice in the CON, CON+CS, HF+Apoe -/- , and HF+Apoe -/- +CS groups was observed to be altered by chronic stress.

Article Snippet: Transverse sections of adrenal glands were incubated with SGLT1 Polyclonal Antibody (1:200, bs-1128R; BIOSS, Beijing, China) and SGLT2 Polyclonal Antibody (1:200, 24654-1-AP; Proteintech, Rosemont, IL, USA).

Techniques: Labeling, Staining

Journal: PeerJ

Article Title: Adrenal SGLT1 or SGLT2 as predictors of atherosclerosis under chronic stress based on a computer algorithm

doi: 10.7717/peerj.15647

Figure Lengend Snippet: (A) The distribution of SGLT1 was observed under low and high magnification, and the differences in SGLT1 expression level in the adrenal gland of mice in the four groups were analyzed by semi-quantitative methods. (B) The distribution of SGLT2 was observed under high and low magnification in the adrenal gland, and the differences in expression between the four groups were analyzed. (C) Edema, glycogen content, and reticulocyte fiber breakage were positively correlated with the expression levels of SGLT1 and SGLT2 in the adrenal gland, respectively.* P < 0.05, ** P < 0.01, *** P < 0.001; ns =no significance.

Article Snippet: Transverse sections of adrenal glands were incubated with SGLT1 Polyclonal Antibody (1:200, bs-1128R; BIOSS, Beijing, China) and SGLT2 Polyclonal Antibody (1:200, 24654-1-AP; Proteintech, Rosemont, IL, USA).

Techniques: Expressing

Journal: PeerJ

Article Title: Adrenal SGLT1 or SGLT2 as predictors of atherosclerosis under chronic stress based on a computer algorithm

doi: 10.7717/peerj.15647

Figure Lengend Snippet: (A) Best training score of 0.0096775 with an epoch of 3000 after BP-neural network training for predicting intima-media thickness of abdominal aorta from SGLT1 and SGLT2 expression levels. (B) Relativity of 0.9841 with good correlation between input and output quantities. (C–D) Validation of the predicted data and the original values, with only small differences. (E) Best training score of 0.012396 with an epoch of 3000 after training the BP-neural network for predicting the internal diameter of abdominal aorta from SGLT1 and SGLT2 expression levels. (F) Relativity of 0.9805, with a good correlation between input and output volumes. (G–H) Validation of predicted data and original values.

Article Snippet: Transverse sections of adrenal glands were incubated with SGLT1 Polyclonal Antibody (1:200, bs-1128R; BIOSS, Beijing, China) and SGLT2 Polyclonal Antibody (1:200, 24654-1-AP; Proteintech, Rosemont, IL, USA).

Techniques: Expressing, Biomarker Discovery

Journal: PeerJ

Article Title: Adrenal SGLT1 or SGLT2 as predictors of atherosclerosis under chronic stress based on a computer algorithm

doi: 10.7717/peerj.15647

Figure Lengend Snippet: (A) The ROC curves showed that the expression level of SGLT1 in the adrenal gland sensitively and specifically predicted adrenal edema, broken reticular fibers, and glycogen content. (B) The ROC curves showed that the expression level of SGLT2 in the adrenal gland sensitively and specifically predicted adrenal edema, broken reticular fibers, and glycogen content.

Article Snippet: Transverse sections of adrenal glands were incubated with SGLT1 Polyclonal Antibody (1:200, bs-1128R; BIOSS, Beijing, China) and SGLT2 Polyclonal Antibody (1:200, 24654-1-AP; Proteintech, Rosemont, IL, USA).

Techniques: Expressing

Journal: PeerJ

Article Title: Adrenal SGLT1 or SGLT2 as predictors of atherosclerosis under chronic stress based on a computer algorithm

doi: 10.7717/peerj.15647

Figure Lengend Snippet: (A) The predicted value of SGLT1 and SGLT2 expression levels on adrenal edema was 0.9596 with a mean error of 3.31% by the SVM method. (B) The predicted value of SGLT1/2 expression levels for broken reticular fibers was 0.8246 with a mean error of 1.81%. (C) The predicted value of SGLT1/2 expression levels for glycogen content was 0.9478 with a mean error of 2.15%.

Article Snippet: Transverse sections of adrenal glands were incubated with SGLT1 Polyclonal Antibody (1:200, bs-1128R; BIOSS, Beijing, China) and SGLT2 Polyclonal Antibody (1:200, 24654-1-AP; Proteintech, Rosemont, IL, USA).

Techniques: Expressing

Journal: PeerJ

Article Title: Adrenal SGLT1 or SGLT2 as predictors of atherosclerosis under chronic stress based on a computer algorithm

doi: 10.7717/peerj.15647

Figure Lengend Snippet: (A–B) Best training score of 0.018092 at epoch 2999 and relativity of 0.96057 of BP-neural network for predicting adrenal edema from SGLT1 and SGLT2 expression levels after training. (C–D) Validation of the predicted values with the original values revealed only small differences. (E–F) A high-risk warning indicator for adrenal edema derived from SGLT1 and SGLT2 expression levels was found by an interpolation algorithm and presented with a three-dimensional (3D) stereogram of the warning range.

Article Snippet: Transverse sections of adrenal glands were incubated with SGLT1 Polyclonal Antibody (1:200, bs-1128R; BIOSS, Beijing, China) and SGLT2 Polyclonal Antibody (1:200, 24654-1-AP; Proteintech, Rosemont, IL, USA).

Techniques: Expressing, Biomarker Discovery, Derivative Assay

Journal: PeerJ

Article Title: Adrenal SGLT1 or SGLT2 as predictors of atherosclerosis under chronic stress based on a computer algorithm

doi: 10.7717/peerj.15647

Figure Lengend Snippet: (A–B) The optimal training score for SGLT1 and SGLT2 to predict adrenal reticular fiber breakage was 0.01571 at epoch 3000 with relativity of 0.96383. (C–D) The predicted data were verified against the original values and a significant difference was found between the two. (E–F) High-risk warning indicators for adrenal reticular fiber breakage were determined from SGLT1 and SGLT2 expression levels, and the warning range was presented with a 3D stereogram.

Article Snippet: Transverse sections of adrenal glands were incubated with SGLT1 Polyclonal Antibody (1:200, bs-1128R; BIOSS, Beijing, China) and SGLT2 Polyclonal Antibody (1:200, 24654-1-AP; Proteintech, Rosemont, IL, USA).

Techniques: Expressing

Journal: PeerJ

Article Title: Adrenal SGLT1 or SGLT2 as predictors of atherosclerosis under chronic stress based on a computer algorithm

doi: 10.7717/peerj.15647

Figure Lengend Snippet: (A–B) The best training score was 0.013784 at epoch 3000 with a relativity of 0.97073. (C–D) Validation revealed only small differences between the predicted values and the original values. (E–F) High-risk warning indicators for adrenal gland glycogen content were determined from SGLT1 and SGLT2 expression levels, and the warning range was presented with a 3D stereogram.

Article Snippet: Transverse sections of adrenal glands were incubated with SGLT1 Polyclonal Antibody (1:200, bs-1128R; BIOSS, Beijing, China) and SGLT2 Polyclonal Antibody (1:200, 24654-1-AP; Proteintech, Rosemont, IL, USA).

Techniques: Biomarker Discovery, Expressing

Journal: Scientific reports

Article Title: Concise and Stereodivergent Synthesis of Carbasugars Reveals Unexpected Structure-Activity Relationship (SAR) of SGLT2 Inhibition.

doi: 10.1038/s41598-017-05895-9

Figure Lengend Snippet: Figure 1. Current SGLT2 inhibitors & carbasugar SGLT2 inhibitors.

Article Snippet: SGLT1- or SGLT2expressing cells (S1 or S2) were developed by transfecting, respectively, SLC5A1 (NM_000343) or SLC5A2 (NM_003041) human cDNA clone (Origene Technologies Inc., Maryland, USA) into COS-7 cells (monkey fibroblast-like kidney cells, CRL-1651, ATCC, Manassas, USA).

Techniques:

Journal: Scientific reports

Article Title: Concise and Stereodivergent Synthesis of Carbasugars Reveals Unexpected Structure-Activity Relationship (SAR) of SGLT2 Inhibition.

doi: 10.1038/s41598-017-05895-9

Figure Lengend Snippet: Figure 7. IC50 values for the novel analogues of dapagliflozin towards SGLT1 and SGLT2.

Article Snippet: SGLT1- or SGLT2expressing cells (S1 or S2) were developed by transfecting, respectively, SLC5A1 (NM_000343) or SLC5A2 (NM_003041) human cDNA clone (Origene Technologies Inc., Maryland, USA) into COS-7 cells (monkey fibroblast-like kidney cells, CRL-1651, ATCC, Manassas, USA).

Techniques: Analogues

Journal: Journal of the American Society of Nephrology

Article Title: Empagliflozin Inhibits Proximal Tubule NHE3 Activity, Preserves GFR, and Restores Euvolemia in Nondiabetic Rats with Induced Heart Failure

doi: 10.1681/asn.2020071029

Figure Lengend Snippet: Figure 3. SGLT2 is overexpressed in the PT of nondiabetic HF rats. (A) Representative immunohistochemical staining of SGLT2 in the PT of sham and HF rats treated with empagliflozin (EMPA) or untreated. Scale bar: 50 mm. (B) Representative immunoblots from SDS-PAGE of renal cortical proteins isolated from sham and HF rats treated with EMPA or untreated and probed with antibodies against SGLT2 and b-actin. (C) Graphic representation of the relative levels of SGLT2 protein abundance in the renal cortex of the four groups of rats. (D) Graphic representation of the relative mRNA expression of SGLT2 in the renal cortex of sham and HF rats treated with EMPA or untreated. The levels of SGLT2 mRNA were measured using quantitative PCR, and GAPDH (glyceraldehyde 3-phosphate dehydroge- nase) was used as an internal control. The values represent individual measurements and the means 6 SEM (Supplemental Material). *P50.01 versus sham; **P50.006 versus sham; ***P,0.001 versus sham; †††P,0.001 versus sham 1 EMPA; ##P50.008 versus HF; ###P,0.001 versus HF.

Article Snippet: The separated proteins were transferred from the gel to a polyvinylidene difluoride membrane (Immobilon-P; Merck Millipore, Bedford, MA) at 350 mA for 8–10 hours at 4 C with a TE 62 Transfer Cooled Unit (GE HealthCare, Piscataway, NJ) and stained with Ponceau S. Then, the polyvinylidene difluoride membranes were incubated with blocking solution (5% nonfat dry milk and 0.1% Tween 20 in PBS, pH 7.4) for 1 hour and overnight (4 C) with specific primary antibodies: a polyclonal antibody against SGLT2 (1:1000, NBP1–92384; Novus Biologicals, Centennial, CO); a mousemAb againstNHE3, clone 3H3 (1:1000; PeterAronson, Yale University, New Haven, CT)22; a phosphospecific mAb against NHE3, PS552-NHE3, clone 14D522 (1:1000; Santa Cruz Biotech, Dallas, TX); or a polyclonal antibody against b-actin (1:5000, ab8227; Abcam).

Techniques: Immunohistochemical staining, Staining, Western Blot, SDS Page, Isolation, Quantitative Proteomics, Expressing, Real-time Polymerase Chain Reaction, Control

Journal: The Korean Journal of Physiology & Pharmacology : Official Journal of the Korean Physiological Society and the Korean Society of Pharmacology

Article Title: Pharmacological evaluation of HM41322, a novel SGLT1/2 dual inhibitor, in vitro and in vivo

doi: 10.4196/kjpp.2019.23.1.55

Figure Lengend Snippet: [ 14 C]-AMG uptake by HEK293 cells overexpressing human SGLT1 (A) and SGLT2 (B). Data are expressed as the mean±S.E.M. of four independent experiments.

Article Snippet: Human SGLT1 and human SGLT2 cDNA were purchased from OriGene (Rockville, MD, USA).

Techniques:

Journal: Cells

Article Title: Canagliflozin, an Inhibitor of the Na + -Coupled D-Glucose Cotransporter, SGLT2, Inhibits Astrocyte Swelling and Brain Swelling in Cerebral Ischemia.

doi: 10.3390/cells12182221

Figure Lengend Snippet: Figure 2. SGLT2 is upregulated in astrocyte cell bodies and astrocyte endfeet following cerebral ischemia. (a–e) Double immunolabeling for SGLT2 (green) and NeuN (a,b) or GFAP (c–e) (red) in contralateral (Contra) and ipsilateral (Ipsi) brain sections from mice post-MCAo/R (2/6 h); merged images are shown in a, b, d, e; individual labelings are shown in c, d—inset, e—inset; the images shown are representative of findings in 3 mice; scale bars, 25 µm except insert, 10 µm.

Article Snippet: The immunoblot of the kidney lysate showed a pair of bands, ~60 and ~65 kDa, with a third band, ~26 kDa, which were absent or minimal in lung lysate and were blocked by pre-absorption of the anti-SGLT2 antibody with the recombinant SGLT2 antigen peptide (#NBP1-92384PEP; Novus Biologicals) (Figure S3a).

Techniques: Immunolabeling

Journal: Cells

Article Title: Canagliflozin, an Inhibitor of the Na + -Coupled D-Glucose Cotransporter, SGLT2, Inhibits Astrocyte Swelling and Brain Swelling in Cerebral Ischemia.

doi: 10.3390/cells12182221

Figure Lengend Snippet: Figure 3. Slc5a2 mRNA and SGLT2 protein are upregulated in astrocytes following cerebral ischemia. (a–c) Images (a) and quantification (b,c) of RNAscope for Slc5a2 (red) and Aqp4 (green) mRNA in contralateral (Contra) and ipsilateral (Ipsi) brain sections from mice post-MCAo/R (2/6 h); scale bar, 25 µm; data from 3 mice per group; **, p < 0.01. (d) qPCR for Slc5a2 mRNA in astrocytes and neurons isolated from the contralateral (Contra) and ipsilateral (Ipsi) MCA territory of mice following MCAo/R (2/6 h) and uninjured controls (naïve), expressed as fold-change; 5 mice per group; *, p < 0.05. (e) Quantitative immunohistochemistry for SGLT2 expressed within regions of interest determined by NeuN; data from 7 mice per group; ns, not significant. (f,g) Immunoblot (f) and quantification of immunoblot (g) for SGLT2 in astrocytes isolated from the contralateral (Contra) and ipsilateral (Ipsi) MCA territory of mice following MCAo/R (2/6 h) and uninjured controls (naïve), expressed as fold-change; PC, positive control (kidney lysate); β-actin used as a loading control; 3 mice per group.

Article Snippet: The immunoblot of the kidney lysate showed a pair of bands, ~60 and ~65 kDa, with a third band, ~26 kDa, which were absent or minimal in lung lysate and were blocked by pre-absorption of the anti-SGLT2 antibody with the recombinant SGLT2 antigen peptide (#NBP1-92384PEP; Novus Biologicals) (Figure S3a).

Techniques: RNAscope, Isolation, Immunohistochemistry, Western Blot, Positive Control, Control

Journal: Cells

Article Title: Canagliflozin, an Inhibitor of the Na + -Coupled D-Glucose Cotransporter, SGLT2, Inhibits Astrocyte Swelling and Brain Swelling in Cerebral Ischemia.

doi: 10.3390/cells12182221

Figure Lengend Snippet: Figure 4. D-glucose-induced Na+ influx in post-ischemic astrocytes is inhibited by canagliflozin and by the astrocyte-specific deletion of Slc5a2/SGLT2. (a) Live cell image of a tdTomato-expressing astrocyte in an ex vivo brain slice from an MCAo/R (2/6 h) mouse, following incubation with ING-2; scale bar, 25 µm. (b–e) Time course of Na+ influx with step change in D-glucose from 2 to 10 mM in ipsilateral (Ipsi) vs. contralateral (Contra) astrocytes (b); in ipsilateral astrocytes without and with pre-incubation with canagliflozin (5 µM) (c); in ipsilateral astrocytes without and with the addition of canagliflozin (5 µM) at the time indicated by the bar (d); in ipsilateral astrocytes from Ast-Slc5a2WT

Article Snippet: The immunoblot of the kidney lysate showed a pair of bands, ~60 and ~65 kDa, with a third band, ~26 kDa, which were absent or minimal in lung lysate and were blocked by pre-absorption of the anti-SGLT2 antibody with the recombinant SGLT2 antigen peptide (#NBP1-92384PEP; Novus Biologicals) (Figure S3a).

Techniques: Expressing, Ex Vivo, Slice Preparation, Incubation