cometadata/datacite-titles-descriptions-related-identifiers huggingface dataset:
DOI: 10.25405/data.ncl.22227079.v1
title: MolMet Figure 3: Lower GKRP and GK protein levels in human NAFLD biopsies of the rs1260326-TT genotype and in the P446L mouse.

description: A-I) Human NAFLD biopsies genotyped for GCKR rs1260326 variant (CC n=23; TT n=17) and scored as simple steatosis (NAFL) or NASH were stained for GKRP or GK and nuclear (N-H) and cytoplasmic (C-H) H-scores were determined. (A) representative images. (B-D) Lower GKRP N-H scores by TT genotype for entire cohort (B) or by -/+ NASH diagnosis (C), lower C-H scores (D) (E-G) Lower GK N-H scores by TT genotype for entire cohort (E) or by -/+ NASH diagnosis (F) and lower C-H scores (G) H) Correlation of GK N-H vs GKRP N-H, r2 0.34, P&lt;0.0001 for entire data set. I) Lower GK (N-H) / GKRP (N-H) ratio by TT genotype, # P&lt; 0.0003. J-M) GKRP P446L mouse aged 14-wk (regular diet, n=4,4, PP, LL) or 28-wk (on HFHSD for last 20-wk, n=10,10, PP, LL): Liver <em>Gckr</em> and <em>Gck</em> mRNA normalized to RplpO (J); GKRP and GK protein (K-L) and GK enzyme activity (M). J) Similar <em>Gckr</em> mRNA by genotype but lower <em>Gck</em> mRNA in 28-wk mice by LL genotype. K-L) Lower GKRP and GK immunoactive protein (n=4-8) and lower GK activity (M, n=4,4,10,10,10) by LL genotype. #P&lt;0.05, LL vs PP; *P&lt;0.05 28-wk vs 14 wk. N-R) Comparative GKRP and GK immunostaining of human liver biopsies (CC,TT: n=7,6) and P446L mouse liver (PP, PL, LL: n=7,4,5) showing representative images for mouse (N) and the N-H and C-H scores. O) Lower GKRP N-H scores for human compared with mouse ($) liver and by TT and LL genotypes (#) and negligible cytoplasmic H-scores. P) Lower GK N-H and C-H for human compared with mouse liver ($) and lower N-H by TT (human) or LL (mouse) genotype (#). Q) Correlation of GK and GKRP N-H scores. R) Relative distribution of GK in nucleus and cytoplasm (N-H/C-H ratio) showing lower nuclear GK sequestration by TT and LL genotypes. #P&lt;0.05 by genotype; $P&lt;0.05 human vs. mouse.
DOI: 10.25405/data.ncl.22227079
title: MolMet Figure 3: Lower GKRP and GK protein levels in human NAFLD biopsies of the rs1260326-TT genotype and in the P446L mouse.

description: A-I) Human NAFLD biopsies genotyped for GCKR rs1260326 variant (CC n=23; TT n=17) and scored as simple steatosis (NAFL) or NASH were stained for GKRP or GK and nuclear (N-H) and cytoplasmic (C-H) H-scores were determined. (A) representative images. (B-D) Lower GKRP N-H scores by TT genotype for entire cohort (B) or by -/+ NASH diagnosis (C), lower C-H scores (D) (E-G) Lower GK N-H scores by TT genotype for entire cohort (E) or by -/+ NASH diagnosis (F) and lower C-H scores (G) H) Correlation of GK N-H vs GKRP N-H, r2 0.34, P&lt;0.0001 for entire data set. I) Lower GK (N-H) / GKRP (N-H) ratio by TT genotype, # P&lt; 0.0003. J-M) GKRP P446L mouse aged 14-wk (regular diet, n=4,4, PP, LL) or 28-wk (on HFHSD for last 20-wk, n=10,10, PP, LL): Liver <em>Gckr</em> and <em>Gck</em> mRNA normalized to RplpO (J); GKRP and GK protein (K-L) and GK enzyme activity (M). J) Similar <em>Gckr</em> mRNA by genotype but lower <em>Gck</em> mRNA in 28-wk mice by LL genotype. K-L) Lower GKRP and GK immunoactive protein (n=4-8) and lower GK activity (M, n=4,4,10,10,10) by LL genotype. #P&lt;0.05, LL vs PP; *P&lt;0.05 28-wk vs 14 wk. N-R) Comparative GKRP and GK immunostaining of human liver biopsies (CC,TT: n=7,6) and P446L mouse liver (PP, PL, LL: n=7,4,5) showing representative images for mouse (N) and the N-H and C-H scores. O) Lower GKRP N-H scores for human compared with mouse ($) liver and by TT and LL genotypes (#) and negligible cytoplasmic H-scores. P) Lower GK N-H and C-H for human compared with mouse liver ($) and lower N-H by TT (human) or LL (mouse) genotype (#). Q) Correlation of GK and GKRP N-H scores. R) Relative distribution of GK in nucleus and cytoplasm (N-H/C-H ratio) showing lower nuclear GK sequestration by TT and LL genotypes. #P&lt;0.05 by genotype; $P&lt;0.05 human vs. mouse.
DOI: 10.25405/data.ncl.22227082
title: MolMet Figure 4: Blood metabolites and liver transcriptome in the P446L mouse on regular diet.

description: A) Male mice aged 14 wk: body weight, free-feeding blood glucose, plasma insulin and plasma triglyceride; n=6-9. B) Female mice aged 14 wk: body weight; plasma triglycerides; liver triglycerides, n=15-17 and glucose tolerance, n=7,7. C) Heat map of differentially expressed genes (DEGs, padjust &lt; 0.05) in livers of male mice by LL genotype and the direction of the log fold change (LFC) of expression values. D) Top pathways from the Ingenuity Pathway Analysis showing corresponding P value and Z scores. E) Venn diagram showing numbers of common genes that are significantly differentially expressed (upregulation, red; down-regulation green) in male mice by LL genotype and are differentially expressed in female PP vs male PP mice. F) Gene counts for P446L male and female mice for common genes from E). G) Box plot of gene counts from RNA-sequencing for 12 <em>Mup</em> genes showing lower expression in male LL mice and in female PP and LL mice; n=4,4,5,5,(M-P, M-L, F-P, F-L).
DOI: 10.25405/data.ncl.22227082.v1
title: MolMet Figure 4: Blood metabolites and liver transcriptome in the P446L mouse on regular diet.

description: A) Male mice aged 14 wk: body weight, free-feeding blood glucose, plasma insulin and plasma triglyceride; n=6-9. B) Female mice aged 14 wk: body weight; plasma triglycerides; liver triglycerides, n=15-17 and glucose tolerance, n=7,7. C) Heat map of differentially expressed genes (DEGs, padjust &lt; 0.05) in livers of male mice by LL genotype and the direction of the log fold change (LFC) of expression values. D) Top pathways from the Ingenuity Pathway Analysis showing corresponding P value and Z scores. E) Venn diagram showing numbers of common genes that are significantly differentially expressed (upregulation, red; down-regulation green) in male mice by LL genotype and are differentially expressed in female PP vs male PP mice. F) Gene counts for P446L male and female mice for common genes from E). G) Box plot of gene counts from RNA-sequencing for 12 <em>Mup</em> genes showing lower expression in male LL mice and in female PP and LL mice; n=4,4,5,5,(M-P, M-L, F-P, F-L).
DOI: 10.25405/data.ncl.22227088
title: MolMet Figure 5: Functional characterization of glucose metabolism and adaptive response to substrate challenge in hepatocytes from P446L mice.

description: A-D) GKRP and GK immunoactivity and enzyme activity after 24h-culture of hepatocytes isolated from male P446L mice (PP and LL genotypes, aged 8-12 wk). (C) Representative immunoblots: protein loading GK 30 μg; GKRP 10 μg (446PP); 35 μg (446LL); D. Glucokinase activity munits/mg protein (n=4-5). E-F) Glucose phosphorylation ([2-<sup>3</sup>H]glucose, E) and glycolysis ([2-<sup>3</sup>H]glucose, F) is similar at 5mM glucose but lower by LL genotype at high glucose, (n=5,5). G) Metabolic scheme showing metabolism of xylitol and fructose bypass the GK reaction and raises glycerol 3-P (G3P); whereas pharmacological activation of GK with PF-04991532 (PF) or inhibition of glucose 6-phosphatase (G6pc) with S4048 (S4) raises glucose 6-P (G6P). Note that fructose at micromolar concentrations also causes activation of GK via elevation in fructose 1-P and thereby elevation in G6P. However at millimolar concentrations the increase in triose phosphates (G3P) is consequent to flux through ketohexokinase and aldolase B reactions. H-K) Hepatocyte incubations with glucose (15mM or 25mM) -/+ the GK activator PF-04991532 (PF, 10μM) or inhibitor of glucose 6-phosphatase (S4048, 1 μM) or xylitol (Xyl, 2mM) or fructose (Fru, 10mM) were for 1h for determination of cell ATP (H), glucose 6-P (I), glycerol 3-P (J) expressed as nmol/mg protein or ATP normalized to respective 5mM (K), n=5,5. L-S) Hepatocyte incubations for RNA extraction and RT-qPCR analysis were for 4h. (L) Basal mRNA levels of genes indicated normalized to Gpd2 mRNA. (M-S) mRNA levels of the genes indicated normalised to the respective 5mM glucose control, n=3,4; # P&lt;0.05 LL vs PP; * P&lt;0.05 relative 5mM glucose control.
DOI: 10.25405/data.ncl.22227088.v1
title: MolMet Figure 5: Functional characterization of glucose metabolism and adaptive response to substrate challenge in hepatocytes from P446L mice.

description: A-D) GKRP and GK immunoactivity and enzyme activity after 24h-culture of hepatocytes isolated from male P446L mice (PP and LL genotypes, aged 8-12 wk). (C) Representative immunoblots: protein loading GK 30 μg; GKRP 10 μg (446PP); 35 μg (446LL); D. Glucokinase activity munits/mg protein (n=4-5). E-F) Glucose phosphorylation ([2-<sup>3</sup>H]glucose, E) and glycolysis ([2-<sup>3</sup>H]glucose, F) is similar at 5mM glucose but lower by LL genotype at high glucose, (n=5,5). G) Metabolic scheme showing metabolism of xylitol and fructose bypass the GK reaction and raises glycerol 3-P (G3P); whereas pharmacological activation of GK with PF-04991532 (PF) or inhibition of glucose 6-phosphatase (G6pc) with S4048 (S4) raises glucose 6-P (G6P). Note that fructose at micromolar concentrations also causes activation of GK via elevation in fructose 1-P and thereby elevation in G6P. However at millimolar concentrations the increase in triose phosphates (G3P) is consequent to flux through ketohexokinase and aldolase B reactions. H-K) Hepatocyte incubations with glucose (15mM or 25mM) -/+ the GK activator PF-04991532 (PF, 10μM) or inhibitor of glucose 6-phosphatase (S4048, 1 μM) or xylitol (Xyl, 2mM) or fructose (Fru, 10mM) were for 1h for determination of cell ATP (H), glucose 6-P (I), glycerol 3-P (J) expressed as nmol/mg protein or ATP normalized to respective 5mM (K), n=5,5. L-S) Hepatocyte incubations for RNA extraction and RT-qPCR analysis were for 4h. (L) Basal mRNA levels of genes indicated normalized to Gpd2 mRNA. (M-S) mRNA levels of the genes indicated normalised to the respective 5mM glucose control, n=3,4; # P&lt;0.05 LL vs PP; * P&lt;0.05 relative 5mM glucose control.
DOI: 10.25405/data.ncl.22227091
title: MolMet Figure 6: Higher blood cholesterol and lower fasting blood glucose and insulin in P446L mice on high energy diets.

description: A-F) Chronic study of P446L mice on HFD: A. Experimental design. B) Body weight, liver weight and epididymis fat pad weight, (n=7,7). C) No difference in free-feeding blood glucose and insulin (21 wk, n=7,7) or glucose tolerance (4-wk, GTT, n=9,9). D) Higher blood cholesterol and liver triglyceride by LL genotype but no difference in blood triglyceride or liver cholesterol (n=6-7). E) Representative hematoxylin-eosin images showing microvesicular steatosis in LL-genotype. F) Histopathology scores for steatosis, microvesicular steatosis, lipogranulomas, lobular Inflammation, portal Inflammation and fibrosis showing higher microvesicular steatosis by LL genotype (n=7,7). G-L) Chronic study of P446L mice on HFHSD: G. Experimental design. H. Body weight, liver weight and higher epididymal fat pad wt by LL genotype, n=11-13. I) Free feeding blood glucose and insulin and glucose tolerance test (GTT), n=11-14. J) Lower post-prandial blood glucose (2h food withdrawal) and insulin and inorganic phosphate (Pi) at cull, n=10-14. K) Higher blood cholesterol in LL genotype but no difference in blood triglycerides or liver triglycerides and cholesterol, n=10-14. L) Histopathology scores showing higher lipogranuloma and lower fibrosis scores by LL genotype, n=14,13. M) Representative hematoxylin-eosin images showing lipogranulomas and Sirius red fast green (SRFG) for fibrosis staining. Categorical data statistical differences was by Chi Square test; other analysis was by t-test #P&lt; 0.05; ##P&lt; 0.01; ### P&lt;0.001 for genotype effect.
DOI: 10.25405/data.ncl.22227091.v1
title: MolMet Figure 6: Higher blood cholesterol and lower fasting blood glucose and insulin in P446L mice on high energy diets.

description: A-F) Chronic study of P446L mice on HFD: A. Experimental design. B) Body weight, liver weight and epididymis fat pad weight, (n=7,7). C) No difference in free-feeding blood glucose and insulin (21 wk, n=7,7) or glucose tolerance (4-wk, GTT, n=9,9). D) Higher blood cholesterol and liver triglyceride by LL genotype but no difference in blood triglyceride or liver cholesterol (n=6-7). E) Representative hematoxylin-eosin images showing microvesicular steatosis in LL-genotype. F) Histopathology scores for steatosis, microvesicular steatosis, lipogranulomas, lobular Inflammation, portal Inflammation and fibrosis showing higher microvesicular steatosis by LL genotype (n=7,7). G-L) Chronic study of P446L mice on HFHSD: G. Experimental design. H. Body weight, liver weight and higher epididymal fat pad wt by LL genotype, n=11-13. I) Free feeding blood glucose and insulin and glucose tolerance test (GTT), n=11-14. J) Lower post-prandial blood glucose (2h food withdrawal) and insulin and inorganic phosphate (Pi) at cull, n=10-14. K) Higher blood cholesterol in LL genotype but no difference in blood triglycerides or liver triglycerides and cholesterol, n=10-14. L) Histopathology scores showing higher lipogranuloma and lower fibrosis scores by LL genotype, n=14,13. M) Representative hematoxylin-eosin images showing lipogranulomas and Sirius red fast green (SRFG) for fibrosis staining. Categorical data statistical differences was by Chi Square test; other analysis was by t-test #P&lt; 0.05; ##P&lt; 0.01; ### P&lt;0.001 for genotype effect.
DOI: 10.25405/data.ncl.22227097.v1
title: MolMet Figure 7: Liver transcriptome analysis in GKRP^P446&gt;L mice on the HFHSD (HD).

description: A) Heat map for unsupervised clustering of P446L livers of PP and LL genotypes on regular diet (P-RD, P-HD, n=4,4) or HFHSD (P-HD, L-HD, n=9,10). B) Ingenuity pathway analysis showing selection of top significantly enriched canonical pathways ranked by P-adjust. C) Venn diagram for converse gene regulation by diet (P-RD x L-HD) and genotype (P-HD x L-HD). Red up-regulation, blue down-regulation. D) Heat map for selected DEGs from groups in C. E) RT-qPCR validation of DEGs (n=4,4,10,12) and comparison with RNA-seq counts (n=4,4,9,10) normalized to P-HD. F) Hmgcr Immunostaining in P446L livers (RD and HD) and cytoplasmic H-scores. n=4,4 (RD); 9,9 (HD). G. Correlation of GK activity and RNA-seq Gck gene counts (n=9,10)/ H) Venn diagram for common genes between the hepatocyte transcriptome (up-regulated by GK and down-regulated by GKRP, Figure 2G) and genes up-regulated by diet (P-RD x P-HD; L-RD x L-HD) I) Correlation coefficient matrix for hepatocyte transcriptome (Figure 2: n=3 hepatocyte preparations,10 conditions) comparing expression of 17-cholesterol linked identified by the IPA analysis on P446L livers and 8 of 20 genes that were up-regulated by GK-overexpression and counter-regulated by GKRP in the hepatocyte study (Figure 2) showing significant (P &lt; 0.05) positive correlations (green) and negative correlations (purple). The Z-score shows up-regulation orange, down regulation grey. J) Correlation coefficient matrix for P446L mouse liver transcriptomes (P-RD, P-RD, P-HD, L-HD, n=27) for the same genes as in (H). The Z-score shows relative expression by experimental group (P-RD, L-RD, P-HD and L-HDL-HD).
DOI: 10.25405/data.ncl.22227097
title: MolMet Figure 7: Liver transcriptome analysis in GKRP^P446&gt;L mice on the HFHSD (HD).

description: A) Heat map for unsupervised clustering of P446L livers of PP and LL genotypes on regular diet (P-RD, P-HD, n=4,4) or HFHSD (P-HD, L-HD, n=9,10). B) Ingenuity pathway analysis showing selection of top significantly enriched canonical pathways ranked by P-adjust. C) Venn diagram for converse gene regulation by diet (P-RD x L-HD) and genotype (P-HD x L-HD). Red up-regulation, blue down-regulation. D) Heat map for selected DEGs from groups in C. E) RT-qPCR validation of DEGs (n=4,4,10,12) and comparison with RNA-seq counts (n=4,4,9,10) normalized to P-HD. F) Hmgcr Immunostaining in P446L livers (RD and HD) and cytoplasmic H-scores. n=4,4 (RD); 9,9 (HD). G. Correlation of GK activity and RNA-seq Gck gene counts (n=9,10)/ H) Venn diagram for common genes between the hepatocyte transcriptome (up-regulated by GK and down-regulated by GKRP, Figure 2G) and genes up-regulated by diet (P-RD x P-HD; L-RD x L-HD) I) Correlation coefficient matrix for hepatocyte transcriptome (Figure 2: n=3 hepatocyte preparations,10 conditions) comparing expression of 17-cholesterol linked identified by the IPA analysis on P446L livers and 8 of 20 genes that were up-regulated by GK-overexpression and counter-regulated by GKRP in the hepatocyte study (Figure 2) showing significant (P &lt; 0.05) positive correlations (green) and negative correlations (purple). The Z-score shows up-regulation orange, down regulation grey. J) Correlation coefficient matrix for P446L mouse liver transcriptomes (P-RD, P-RD, P-HD, L-HD, n=27) for the same genes as in (H). The Z-score shows relative expression by experimental group (P-RD, L-RD, P-HD and L-HDL-HD).
DOI: 10.25405/data.ncl.22227100
title: MolMet Figure 8: Correlation of liver transcripts with phenotypic traits in the P446L mouse on the HFHSD.

description: A) Correlation of phenotypic traits with corresponding gene counts from the liver transcriptome for the P446L mouse (PP,LL, n=9/10) on the HSFSD showing: (i) Blood glucose (BG) and blood insulin (BI) correlated negatively with genes induced by GK excess (<em>Scl2a2</em>, <em>Slc25a47</em>, <em>Rgs16</em>, <em>G6pc</em>, <em>Rorc</em>) in the hepatocyte transcriptome (Figure 2). (ii) Blood glucose and insulin correlated positively with <em>Hmgcr</em> and other genes linked to cholesterol biosynthesis (<em>Msmo1</em>, <em>Insig2</em>, <em>Cyp51a1</em>) and negatively with genes linked to cholesterol excretion (<em>Abcg5</em>, <em>Abcg8</em>). (iii) Inorganic phosphate (Pi) correlated negatively with <em>G6pc</em>; (iv) Blood cholesterol (TC, HDL, LDL) correlated positively with <em>Slc25a47</em> and <em>Slc25a10</em> and negatively with <em>Tm7sf2</em>).
DOI: 10.25405/data.ncl.22227100.v1
title: MolMet Figure 8: Correlation of liver transcripts with phenotypic traits in the P446L mouse on the HFHSD.

description: A) Correlation of phenotypic traits with corresponding gene counts from the liver transcriptome for the P446L mouse (PP,LL, n=9/10) on the HSFSD showing: (i) Blood glucose (BG) and blood insulin (BI) correlated negatively with genes induced by GK excess (<em>Scl2a2</em>, <em>Slc25a47</em>, <em>Rgs16</em>, <em>G6pc</em>, <em>Rorc</em>) in the hepatocyte transcriptome (Figure 2). (ii) Blood glucose and insulin correlated positively with <em>Hmgcr</em> and other genes linked to cholesterol biosynthesis (<em>Msmo1</em>, <em>Insig2</em>, <em>Cyp51a1</em>) and negatively with genes linked to cholesterol excretion (<em>Abcg5</em>, <em>Abcg8</em>). (iii) Inorganic phosphate (Pi) correlated negatively with <em>G6pc</em>; (iv) Blood cholesterol (TC, HDL, LDL) correlated positively with <em>Slc25a47</em> and <em>Slc25a10</em> and negatively with <em>Tm7sf2</em>).
DOI: 10.25405/data.ncl.22227103
title: MolMet Supp 1: Optimization of adenoviral vector titres for comparing human and mouse GKRP.

description: A) Experimental design: hepatocytes isolated from GKRP-deficient mice were cultured in monolayer and transfected with adenoviral vectors for expression of human or mouse GKRP(446P or 446L) at titres of 5 to 30 x 10<sup>6</sup> pfu/ml and cultured for 24h for analysis of mRNA, immunoactivity and immunostaining for GKRP (AG-GKRP) and GK (AG-GK). B) No nuclear GKRP or GK immunostaining in un-transfected GKRP deficient hepatocytes. C-E) Nuclear GKRP and GK staining in transfections with human GKRP (NM_001486.4; NP_001477.2) h-446P/L at titres of 10,20,30 x 10<sup>6</sup> pfu/ml. (C) GKRP immunoreactivity compared with untransfected GKRP<sup>+/-</sup> hepatocytes (Het). (D) Representative images of nuclear staining of GKRP and GK. E) Maximum nuclear sequestration of GKRP at the lowest titres of 10x10<sup>6</sup> pfu/ml, measured from the nuclear / cytoplasmic (N/C) intensity ratio, * P&lt; 0.02 vs 10x10<sup>6</sup> pfu/ml. F-G) Exclusive cytoplasmic staining for GKRP and GK in hepatocytes transfected mouse Gckr-Transcript-2 (NM_144909.2; NP_659158.1) 446P/L. (F) GKRP immunoreactivity (mouse transcript-2) compared with untransfected GKRP<sup>+/-</sup> hepatocytes (Het). (G) Immunostaining showing exclusive cytoplasmic staining for GKRP and GK. H) Nuclear GKRP and GK staining in transfections with mouse GKRP-Transcript-1 (NM_001374741.1; NP_001361670.1) 446P/L at 5x10<sup>6</sup> and 10x10<sup>6</sup> pfu/ml. I) Lack of nuclear staining in cells transfected with Gckr-X1 (XM-006503882) and Gckr-X2 (XM-006503883). J) Similar cellular transfection efficiency at 5x10<sup>6</sup> and 10x10<sup>6</sup> pfu/ml for human GKRP (hP,hL) or mouse T1 (mP,mL) determined from GKRP stained nuclei % DAPI staining, n=3 hepatocyte preparations. K) Similar Human <em>GCKR</em> or mouse <em>Gckr</em>-T1 transcript levels (normalized to <em>Gapdh</em> mRNA) for 446P and 446L vectors in transfections with human or mouse GKRP:446P or L (at 5 or 10 x 10<sup>6</sup> pfu/ml). Means ± SEM for n=8 (human GKRP), n=7 (mouse GKRP) hepatocyte experiments.
DOI: 10.25405/data.ncl.22227103.v1
title: MolMet Supp 1: Optimization of adenoviral vector titres for comparing human and mouse GKRP.

description: A) Experimental design: hepatocytes isolated from GKRP-deficient mice were cultured in monolayer and transfected with adenoviral vectors for expression of human or mouse GKRP(446P or 446L) at titres of 5 to 30 x 10<sup>6</sup> pfu/ml and cultured for 24h for analysis of mRNA, immunoactivity and immunostaining for GKRP (AG-GKRP) and GK (AG-GK). B) No nuclear GKRP or GK immunostaining in un-transfected GKRP deficient hepatocytes. C-E) Nuclear GKRP and GK staining in transfections with human GKRP (NM_001486.4; NP_001477.2) h-446P/L at titres of 10,20,30 x 10<sup>6</sup> pfu/ml. (C) GKRP immunoreactivity compared with untransfected GKRP<sup>+/-</sup> hepatocytes (Het). (D) Representative images of nuclear staining of GKRP and GK. E) Maximum nuclear sequestration of GKRP at the lowest titres of 10x10<sup>6</sup> pfu/ml, measured from the nuclear / cytoplasmic (N/C) intensity ratio, * P&lt; 0.02 vs 10x10<sup>6</sup> pfu/ml. F-G) Exclusive cytoplasmic staining for GKRP and GK in hepatocytes transfected mouse Gckr-Transcript-2 (NM_144909.2; NP_659158.1) 446P/L. (F) GKRP immunoreactivity (mouse transcript-2) compared with untransfected GKRP<sup>+/-</sup> hepatocytes (Het). (G) Immunostaining showing exclusive cytoplasmic staining for GKRP and GK. H) Nuclear GKRP and GK staining in transfections with mouse GKRP-Transcript-1 (NM_001374741.1; NP_001361670.1) 446P/L at 5x10<sup>6</sup> and 10x10<sup>6</sup> pfu/ml. I) Lack of nuclear staining in cells transfected with Gckr-X1 (XM-006503882) and Gckr-X2 (XM-006503883). J) Similar cellular transfection efficiency at 5x10<sup>6</sup> and 10x10<sup>6</sup> pfu/ml for human GKRP (hP,hL) or mouse T1 (mP,mL) determined from GKRP stained nuclei % DAPI staining, n=3 hepatocyte preparations. K) Similar Human <em>GCKR</em> or mouse <em>Gckr</em>-T1 transcript levels (normalized to <em>Gapdh</em> mRNA) for 446P and 446L vectors in transfections with human or mouse GKRP:446P or L (at 5 or 10 x 10<sup>6</sup> pfu/ml). Means ± SEM for n=8 (human GKRP), n=7 (mouse GKRP) hepatocyte experiments.