Uses of knockout, knockdown, and transgenic models in the studies of glucose transporter 4

Table 3 The transgenic studies using the SLC2A4 mini gene and its promoter for the whole-body expression in mice

Transgenic constructs	Analysis	Observations	Ref.
A 11.5-kb mini gene of human SLC2A4 starts with a 5.3-kb fragment upstream of transcription start and terminates within exon 10 of the gene followed by the bacterial CAT in pHSS6 vector	RNase protection assay and Western blot were used for SLC2A4 mRNA and GLUT4 protein in BAT, WAT, heart and skeleton muscle, respectively	The transgene expression was detected in WAT and BAT, heart and skeleton muscle of mice. Female transgenic mice have higher GLUT4 protein in the adipose tissue and less SLC2A4 mRNA in skeleton muscle than male ones. Transgenic mice have higher GLUT4 protein level in adipose tissue, liver, heart and skeleton muscle than the controls	[43]
The 11.5-kb minigene with the CAT reporter as shown in[43]	Reverse transcription PCR was used to measure SLC2A4 mRNA in cardiac and hindquarter muscle, BAT and WAT. Immunofluorescent test was for GLUT4 translocation	Transgenic mice gained more weight after 15 wk old of age, and have lower blood glucose in both fasting and fed states, lower insulin level in fasting and higher after refeeding, and higher glycogen contents, GLUT4 translocation in cardiac and skeleton muscle than the control mice	[44]
The 11.5 kb minigene with the CAT reporter as shown in[43]	Western blot was used to detect GLUT4 in gastrocnemius muscles	Transgenic mice have lower serum glucose level in both fasting and fed state, higher insulin level during fasting and lower after fed than the control ones	[45]
The 11.5 kb minigene with the CAT reporter as shown in[43]	Western blot was used to detect GLUT4 in the heart	Transgenic mice have higher glucose uptake, glycolysis and glycogen content, and lower insulin-stimulated glycolysis rate and glycogen synthesis in the heart than the control ones. Glucose and fatty acid oxidation remain the same	[46]
The 11.5 kb minigene with the CAT reporter as shown in[43]	Immunofluorescence was used to detect GLUT4 in cardiac myocytes and adipocytes	Transgenic mice have similar body weight, and epididymal adipose tissue weight and adipocyte size as the controls. Transgenic mice have higher levels of triglycerides, β-hydroxybutyrate and free fatty acids, and parametrial fat weight and lower glucose level after an oral glucose challenge and insulin level after an insulin injection than the controls. The insulin-stimulated glucose uptake is impaired in transgenic mice	[47]
A 2.4-kb of 5’ flanking DNA fragment of human SLC2A4 promoter fused with the CAT as a reporter construct	CAT activity assay and RNase protection assay were used to detect promoter activation and mRNA, respectively	In transgenic mice, CAT activity can be detected in the tissues that generally express GLUT4, including BAT and WAT, and smooth, skeleton and cardiac muscle, but not the liver	[42]
A 2.4-kb of 5’ flanking DNA of human SLC2A4 promoter fused to CAT as shown in[42]	Western blot was used to detect GLUT4 in adipose and skeleton muscle tissues	Transgenic mice have slower rise of blood glucose (no difference in glucose and insulin levels) during pentobarbital sodium anesthesia, and higher glucose infusion rate (40% increase) during hyper insulinemic euglycemic clamp than the controls	[48]
A 2.4-kb of 5 flanking DNA of human SLC2A4 promoter fused to CAT as shown in[42]	Only cited previous publications[42]	Transgenic mice have lower blood glucose, higher lactate and β-hydroxybutyrate levels during both fasting and fed states, and better glucose transport in the soleus muscle when fed a high-fat and high-sugar diet than the controls	[49]

BAT: Brown adipose tissue; CAT: Chloramphenicol acetyltransferase; GLUT4: Glucose transporter 4; Ref: References; WAT: White adipose tissue.