Knock-in of diphteria toxin A chain gene at Ins2 locus: effects on islet development and localization of Ins2 expression in the brain

Abstract

We report here knock-in of diphteria toxin A chain (dta) gene at the Ins2 locus, using the strategy previously employed to insert lacZ under control of the Ins2 promoter. Mutant Ins2(dta/+), Ins2(dta/lacZ) or Ins2(lacZ/+) mouse pups were generated by breeding and analyzed to study the effects of toxigenetic beta-cell ablation on islet development and to localize the extrapancreatic Ins2 expression site in the brain. Ins2(dta/+) and Ins2(dta/lacZ) pups developed a severe diabetic ketoacidosis and died rapidly. Histological analysis of their pancreas revealed that beta-cells completely disappeared in their islets as evidenced by loss of lacZ activity or insulin immunonostaining. beta-cell ablation did not alter the size of other islet cell populations which were normal at birth, although the glucagon-cell population was reduced by 85% at embryonic day E12.5. In the brain, comparative analysis of lacZ expression in Ins2(lacZ/+) and Ins2(dta/laZ) mice identified the choroid plexus (CP) as a major Ins2 expression site. This finding was confirmed by RT-PCR analysis of insulin transcripts in RNAs prepared from microdissected wild-type CP. Transcripts for other key beta-cell markers, with the notable exception of Pdx-1, were also found in CP RNAs. These results must revive interest in studies focused on extrapancreatic insulin gene expression.

Efficient gene delivery to pancreatic islets with ultrasonic microbubble destruction technology

“…The insulin 2 gene is expressed in pancreatic beta cells, thymus, choroid plexus, and yolk sac. The insulin 1 gene is expressed only in pancreatic beta cells and scattered cells in the thymus (18)(19)(20)(21)(22). Thus, we used a fragment of the RIP (Ϫ412 to ϩ155 relative to the transcription start site) for our vector construction to facilitate gene targeting to beta cells.…”

Effects of high-fat diet feeding on Znt8-null mice: differences between β-cell and global knockout of Znt8

“…Nevertheless, Ins2 promoter activity was reported in many other extrahypothalamic brain areas. The choroid plexus (30), cortex, caudate putamen, ventral pallidum, substantia nigra, pons (17), and mid-and ventral regions of the brain were all shown to have Ins2 promoter activity (62). Importantly, Znt8 expression was reported in the cortex, as seen in the Allen Mouse Brain Atlas (http://www.brain-map.org) (33).…”

Mutations in gfpt1 and skiv2l2 cause distinct stage-specific defects in larval melanocyte regeneration in zebrafish

“…Additionally, it may also provide an opportunity to understand the cause or cure for human disease resulting from single cell type deficits, such as loss of b cells in autoimmune type I diabetes [12]. Several methods that ablate single cell types have been reported in various vertebrates, including genetic approaches, and use of lasers or chemicals [10,[13][14][15][16]. For example, Lamotte et al [13] used transgenic methods to introduce tissue-specific expression of a toxin to ablate mouse pancreatic b cells.…”

“…Several methods that ablate single cell types have been reported in various vertebrates, including genetic approaches, and use of lasers or chemicals [10,[13][14][15][16]. For example, Lamotte et al [13] used transgenic methods to introduce tissue-specific expression of a toxin to ablate mouse pancreatic b cells. Other examples include exposure to a dermatology laser and incubation with a small molecule, 4-(4-morpholinobutylthio)phenol (MoTP), each of which has been shown to specifically ablate zebrafish larval melanocytes [9,17].…”