Transcriptome profiling of human hypothalamic agouti-related protein and proopiomelanocortin neurons regulating energy homeostasis.

Methodological development and validation of IHC/LCM-Seq for transcriptome profiling in postmortem human brain tissue

“We present and validate a pioneering ‘IHC/LCM-Seq’ method for transcriptome profiling of spatially defined neuronal cell types detected with immunohistochemistry in sections of formaldehyde-fixed human brains.”

Postmortem human brains from 13 adult donors (1 female, 12 males) of European ancestry

“Histological samples from adult human brains (N = 13) were obtained during autopsies conducted at the Department of Pathology and Experimental Cancer Research, Semmelweis University, Budapest, Hungary.”

Transcriptomic analysis of 8 libraries comprising 3 AgRP, 2 POMC, and 3 KP neuron samples; spatial analysis of 2 subjects

“Sequencing of eight libraries—comprising 3 AgRP-, 2 POMC-, and 3 KP neuron transcriptomes—generated a total of 211.8 million raw reads (17.6–33 million per sample).”

Postmortem delay kept below 24 hours; immersion fixation for 24-72 hours

“Postmortem time was kept below 24 h.”

The IHC/LCM-Seq method identified 14,000–16,000 transcripts per cell type in AgRP, POMC, and KP neurons

“Application of IHC/LCM-Seq to male human hypothalami enables the identification of 14,000–16,000 transcripts in agouti-related protein (AgRP) neurons, which drive appetite and energy storage, and proopiomelanocortin (POMC) neurons, which suppress feeding and promote energy expenditure.”

AgRP and POMC neurons exhibit distinct receptor profiles, with AgRP neurons enriched for proinflammatory cytokine receptors and POMC neurons for reproductive hormone and endocannabinoid receptors

“The AgRP neuron transcriptome is rich in receptors for proinflammatory cytokines, metabolic hormones and growth hormone, whereas POMC neurons express reproductive hormone-, glucagon-like peptide-, calcitonin-, and endocannabinoid receptors.”

Significant species-specific differences in neuropeptide and receptor expression were observed between human and mouse AgRP and POMC neurons

“Analysis of neuropeptides with mean expression ≥40 TPM in human or mouse AgRP or POMC neuron transcriptomes, including mouse fed and food-deprived states, revealed pronounced species differences.”

The study used a male-only analytic dataset of European-ancestry adults, which may constrain generalizability to other populations

“We note that the lack of socioeconomic status data and the male-only analytic dataset of European-ancestry adults may constrain the generalizability of the findings to other populations.”