Journal of Pineal Research最新文献

Disruptions in circadian rhythm, partly controlled by the hormone melatonin, increase the risk of type 2 diabetes (T2D). Accordingly, a variant of the gene encoding the melatonin receptor 1B (MTNR1B) is robustly associated with increased risk of T2D. This single-nucleotide polymorphism (SNP; rs10830963; G-allele) is an expression quantitative trait locus (eQTL) in human pancreatic islets, conferring increased expression of MTNR1B, which is thought to perturb pancreatic β-cell function. To understand this pathogenic mechanism in detail, we utilized human induced pluripotent stem cells (hiPSC), derived from individuals with T2D carrying the MTNR1B G-allele. Patient-derived fibroblasts were reprogrammed to hiPSC and single-base genome editing by CRISPR/Cas9 was employed to create isogenic lines of either the C/C or G/G genotypes (nonrisk and risk, respectively). In addition, the human embryonic stem cell (hESC) line (HUES4) was subjected to genome editing to create isogenic lines of either the C/C or G/G genotypes. hiPSC and hESC were differentiated into β-like cells, using a 50-day 2D protocol. Single-base genome editing generated cells with the desired genotype at a success rate of > 90%. Expression of stage-specific markers confirmed differentiation of both hiPSC and hESC into β-cells. MTNR1B mRNA levels were consistently low in differentiated β-cells, precluding quantitative analysis of gene expression. Western blot analyses indicated slightly higher levels of the MTNR1B protein in differentiated β-cells carrying the risk allele, which is in accord with the notion that rs10830963 (G-allele) functions as an eQTL in β-cells. Insulin secretion in response to the combination of high glucose and IBMX was comparable between genotypes, whereas the addition of melatonin appeared to reduce insulin secretion more efficiently in cells carrying the G-allele. While our data suggest elevated MTNR1B protein levels in stem cell-derived β-like cells carrying the risk allele, these cells do not appear to be sufficiently mature to establish rs10830963 as an eQTL at the mRNA level. The observed nominal increase in melatonin sensitivity in G-allele–carrying cells is suggestive of a functional contribution of rs10830963 to β-cell dysfunction; however, this interpretation remains tentative and will require further validation in more mature β-cell models.

Melatonin, an indolamine primarily recognized for regulating circadian rhythms, has also demonstrated notable antitumoral properties. Melatonin induces endoplasmic reticulum (ER) stress, modulates autophagy, and promotes apoptosis in various tumors, including gastric, ovarian, cervical, oral tongue, colorectal, renal, hepatic, and bladder cancer. In placental choriocarcinoma, melatonin reduces cell viability and induces apoptosis by inhibiting autophagy and disrupting the mitochondrial membrane potential. However, its effects on ER stress and the unfolded protein response (UPR) pathway remain unexplored. It is hypothesized here that the proapoptotic effects of melatonin in choriocarcinoma cells occur through the activation of the UPR pathway. The factors implicated in the UPR (PERK, IRE1ɑ, ATF6, GRP78, ATF4, CHOP, P-eIF2α) pathways were evaluated by Western blot, RT-qPCR, and flow cytometry in BeWo (human choriocarcinoma) cells treated with or without melatonin (1 mM). Melatonin significantly increased protein levels of GRP78 (p = 0.0329), IRE1α (p = 0.0394), p-eIF2α (p = 0.0439), ATF4 (p = 0.0267), CHOP (p = 0.0379), Bax and cleaved PARP but did not affect TRAF2 and NFkB protein levels nor XBP1 mRNA splicing. PERK knockdown, via siRNA, prevented the rise in GRP78, p-eIF2α/eIF2α, and ATF4 levels by melatonin. Additionally, melatonin increased early apoptosis in BeWo cells (p = 0.0371) and PERK knockdown increased the susceptibility of BeWo cells to apoptosis when treated with tunicamycin (p = 0.0359), suggesting that ER stress plays a role in BeWo cell survival. This study demonstrates that melatonin activates the PERK-ATF4-P-eIF2α-CHOP pathway and induces early apoptosis in BeWo cells, while PERK deficiency compromises cell survival under ER stress. Our findings suggest that modulating PERK-UPR signaling with melatonin could present a promising therapeutic strategy for cancer, including placental choriocarcinoma.

Circadian rhythmicity plays a crucial role in regulating sleep timing and continuity, but it may be altered in people with insomnia. This study tested whether dim-light melatonin onset (DLMO) timing is associated with sleep difficulties in insomnia. In total, 128 people diagnosed with insomnia disorder were recruited. Participants completed daily sleep diaries and wore an actigraph for up to 14 days before the laboratory visit to estimate mean sleep continuity (e.g., sleep latency, sleep duration) and sleep timing (sleep onset time and wake time from diaries, bedtime from diaries and actigraphy). After a sleep study, participants underwent salivary melatonin collection to estimate DLMO on the following night. Regressions and analyses of variances on tertile groups tested associations between DLMO (clock times and phase angle differences between DLMO and sleep timing) and sleep continuity and timing. There were associations between DLMO and sleep timing, r_(s) = 0.27–0.37, but not with sleep continuity. The phase angle between sleep onset time and DLMO was associated with sleep latency, sleep duration, and sleep efficiency, r_(s) = −0.32 to 0.41. Participants with a longer phase angle between DLMO and sleep onset time (> 3 h; i.e., greater delays) had longer sleep latencies (Mean diff = 43.21 min, SE = 12.99, p = 0.004) and shorter sleep durations (Mean diff = −65.66 min, SE = 20.75, p = 0.006) than participants with a shorter phase angle (< 2 h). Other phase angles (DLMO and mid-sleep, wake time) were not consistently associated with sleep continuity. Melatonin onset timing is associated with sleep timing in insomnia disorder. Larger phase angle differences between sleep onset and DLMO are linked to poorer sleep continuity. These findings highlight the importance of considering circadian alignment and its impact on sleep in understanding the pathophysiology of insomnia and in developing targeted treatment interventions.

In response to Yoshiyasu Takefuji's critique regarding the use of Principal Component Analysis (PCA) and Partial Least Squares Discriminant Analysis (PLS-DA) in the study “Melatonin Repairs the Lipidome of Human Hepatocytes Exposed to Cd and Free Fatty Acid-Induced Lipotoxicity,” we provide a methodological clarification. PCA and PLS-DA are well-established, widely validated tools for exploratory analysis of high-dimensional omics data, including lipidomics data. Although these methods are linear, they are appropriate for capturing systematic and directional variations in complex biological systems, particularly in controlled in vitro models like ours. Our analytical approach integrates PCA and PLS-DA with rigorous statistical testing, data transformations, and biological validation, ensuring robustness and biological relevance of the findings. We reaffirm that these methods represent a standard, reliable practice in lipidomics, and the potential of nonlinear techniques does not diminish the appropriateness or utility of linear multivariate models when applied with scientific rigor.

Rest-activity rhythm and sleep may serve as potential intervention targets for a variety of diseases. However, the underlying biological mechanisms of rest-activity rhythm, sleep, and their proteomic associations with multiple diseases remain largely unexplored. Here, using data from approximately 10 000 participants in the UK Biobank with accelerometer-derived measures and proteomics profiles, we characterized the proteomic signatures of rest-activity and sleep and explored their associations with health outcomes. We found that the proteins associated with rest-activity and sleep were mainly enriched in inflammation, immune response and complement system. Most rest-activity and sleep measures, along with their associated proteomic signatures, were significantly associated with incident diabetes, cardiovascular disease, chronic kidney disease, respiratory diseases, and extended life expectancy. Several proteins, such as ADM and CA14, were observed to mediate multiple associations across distinct rest-activity and sleep measures. The impact of rest-activity and sleep measures on chronic diseases and mortality may be mediated through diverse biological pathways involving multiple proteins. These findings reveal potential mechanisms underlying these complex relationships and provide novel insights for the development of targeted intervention strategies.