Heon-Jin Lee

Oxytocin receptor knockout mice display deficits in the expression of autism-related behaviors

A wealth of studies has implicated oxytocin (Oxt) and its receptors (Oxtr) in the mediation of social behaviors and social memory in rodents. It has been suggested that failures in this system contribute to deficits in social interaction that characterize autism spectrum disorders (ASD). In the current analyses, we investigated the expression of autism-related behaviors in mice that lack the ability to synthesize the oxytocin receptor itself, Oxtr knockout (KO) mice, as compared to their wild-type (WT) littermates. In the visible burrow system, Oxtr KO mice showed robust reductions in frontal approach, huddling, allo-grooming, and flight, with more time spent alone, and in self-grooming, as compared to WT. These results were corroborated in the three-chambered test: unlike WT, Oxtr KO mice failed to spend more time in the side of the test box containing an unfamiliar CD-1 mouse. In the social proximity test, Oxtr KO mice showed clear reductions in nose to nose and anogenital sniff behaviors oriented to an unfamiliar C57BL/6J (B6) mouse. In addition, our study revealed no differences between Oxtr WT and KO genotypes in the occurrence of motor and cognitive stereotyped behaviors. A significant genotype effect was found in the scent marking analysis, with Oxtr KO mice showing a decreased number of scent marks, as compared to WT. Overall, the present data indicate that the profile for Oxtr KO mice, including consistent social deficits, and reduced levels of communication, models multiple components of the ASD phenotype. This article is part of a Special Issue entitled Oxytocin, Vasopressin, and Social Behavior.

MicroRNA-124 regulates osteoclast differentiation.

Osteoclasts are specialized cells for bone-resorption originated from precursors of macrophage/monocyte lineage. The receptor activator of NFκB ligand (RANKL) initiates osteoclast differentiation, in which nuclear factor of activated T cell cytoplasmic 1 (NFATc1) plays a key role as a master transcription factor. In the present report, we show that microRNA-124 (miR-124) regulates osteoclastogenesis of mouse bone marrow macrophages (BMMs) by suppressing NFATc1 expression. On the other hand, synthetic inhibitor that binds specifically to miR-124 enhanced osteoclast differentiation and NFATc1 expression. The overexpression of a constitutively active form of NFATc1 prevented the inhibitory effect of miR-124 on osteoclastogenesis. Finally, miR-124 also affected the proliferation and motility of osteoclast precursors, the latter coinciding with the reduced expression of RhoA and Rac1. These findings not only reveal unprecedented role of miR-124 in osteoclastogenesis but also suggest a novel mode of regulation of NFATc1 in osteoclasts.

Normal Maternal Behavior, but Increased Pup Mortality, in Conditional Oxytocin Receptor Knockout Females

Oxytocin (Oxt) and the Oxt receptor (Oxtr) are implicated in the onset of maternal behavior in a variety of species. Recently, we developed two Oxtr knockout lines: a total body knockout (Oxtr-/-) and a conditional Oxtr knockout (OxtrFB/FB) in which the Oxtr is lacking only in regions of the forebrain, allowing knockout females to potentially nurse and care for their biological offspring. In the current study, we assessed maternal behavior of postpartum OxtrFB/FB females toward their own pups and maternal behavior of virgin Oxtr-/- females toward foster pups and compared knockouts of both lines to wildtype (Oxtr+/+) littermates. We found that both Oxtr-/- and OxtrFB/FB females appear to have largely normal maternal behaviors. However, with first litters, approximately 40% of the OxtrFB/FB knockout dams experienced high pup mortality, compared to fewer than 10% of the Oxtr+/+ dams. We then went on to test whether or not this phenotype occurred in subsequent litters or when the dams were exposed to an environmental disturbance. We found that regardless of the degree of external disturbance, OxtrFB/FB females lost more pups on their first and second litters compared to wildtype females. Possible reasons for higher pup mortality in OxtrFB/FB females are discussed.

Bone marrow oxytocin mediates the anabolic action of estrogen on the skeleton

Background: The mechanism underlying the anabolic effect of estrogen on the skeleton is unclear. Results: We report that estrogen-induced bone formation in mice occurs through oxytocin (OT) produced by osteoblasts in bone marrow. Conclusion: Feed-forward OT release in bone marrow by a rising estrogen level may facilitate rapid skeletal recovery after lactation. Significance: The study highlights a novel mechanism for estrogen action on bone. Estrogen uses two mechanisms to exert its effect on the skeleton: it inhibits bone resorption by osteoclasts and, at higher doses, can stimulate bone formation. Although the antiresorptive action of estrogen arises from the inhibition of the MAPK JNK, the mechanism of its effect on the osteoblast remains unclear. Here, we report that the anabolic action of estrogen in mice occurs, at least in part, through oxytocin (OT) produced by osteoblasts in bone marrow. We show that the absence of OT receptors (OTRs) in OTR−/− osteoblasts or attenuation of OTR expression in silenced cells inhibits estrogen-induced osteoblast differentiation, transcription factor up-regulation, and/or OT production in vitro. In vivo, OTR−/− mice, known to have a bone formation defect, fail to display increases in trabecular bone volume, cortical thickness, and bone formation in response to estrogen. Furthermore, osteoblast-specific Col2.3-Cre+/OTRfl/fl mice, but not TRAP-Cre+/OTRfl/fl mice, mimic the OTR−/− phenotype and also fail to respond to estrogen. These data attribute the phenotype of OTR deficiency to an osteoblastic rather than an osteoclastic defect. Physiologically, feed-forward OT release in bone marrow by a rising estrogen concentration may facilitate rapid skeletal recovery during the latter phases of lactation.

Impairments in the initiation of maternal behavior in oxytocin receptor knockout mice.

Oxytocin (Oxt) acting through its single receptor subtype, the Oxtr, is important for the coordination of physiology and behavior associated with parturition and maternal care. Knockout mouse models have been helpful in exploring the contributions of Oxt to maternal behavior, including total body Oxt knockout (Oxt −/−) mice, forebrain conditional Oxtr knockout (Oxtr FB/FB) mice, and total body Oxtr knockout (Oxtr −/−) mice. Since Oxtr −/− mice are unable to lactate, maternal behavior has only been examined in virgin females, or in dams within a few hours of parturition, and there have been no studies that have examined their anxiety-like and depression-like behavior following parturition. To improve our understanding of how the absence of Oxt signaling affects maternal behavior, mood and anxiety, we designed a study using Oxtr −/− mice that separated nursing behavior from other aspects of maternal care, such as licking and grooming by thelectomizing (i.e. removing the nipples) of Oxtr +/+ mice and sham-thelectomizing Oxtr −/− mice, and pairing both genotypes with a wet nurse. We then measured pup abandonment, maternal behavior, and postpartum anxiety-like and depression-like behaviors. We hypothesized that genetic disruption of the Oxtr would impact maternal care, mood and anxiety. Specifically, we predicted that Oxtr −/− dams would have impaired maternal care and increased anxiety-like and depression-like behaviors in the postpartum period. We found that Oxtr −/− dams had significantly higher levels of pup abandonment compared to controls, which is consistent with previous work in Oxtr FB/FB mice. Interestingly, Oxtr −/− dams that initiated maternal care did not differ from wildtype controls in measures of maternal behavior. We also did not find any evidence of altered anxiety-like or depressive-like behavior in the postpartum period of Oxtr −/− dams. Thus, our data suggest that Oxt lowers the threshold for the initiation of maternal behavior.

MicroRNA-365 regulates NKX2-1, a key mediator of lung cancer

MicroRNAs constitute a class of small noncoding RNAs that play roles in tumorigenesis. We found that NKX2-1 protein levels were generally high in the lung cancer tissues whereas miRNA-365 expression levels were downregulated. Ectopic miR-365 expression decreased NKX2-1 expression in lung cancer cell lines. Transfection of a miR-365 mimic led to reduced proliferation of lung cancer cells; conversely, a miR-365 inhibitor slightly increased cell proliferation. The NKX2-1 overexpression significantly increased the cell proliferation by overcoming the suppressive effect of miR-365. Our data suggest that miR-365 is an important regulator of NKX2-1 and can be a target for lung cancer therapies.

Heightened aggressive behavior in mice with lifelong versus postweaning knockout of the oxytocin receptor.

Previous work implicating the neuropeptide oxytocin (Oxt) in the neural regulation of aggression in males has been limited. However, there are reports of heightened aggression in Oxt knockout and Oxt receptor (Oxtr) knockout male mice when they are born to null mutant mothers; suggesting that intrauterine exposure to Oxt may be important to normal aggression in adulthood. To explore this, we examined aggression in two lines of Oxtr mice, a total knockout (Oxtr-/-), in which the Oxtr gene is absent from the time of conception, and a predominantly forebrain specific knockout (Oxtr FB/FB), in which the Oxtr gene is not excised until approximately 21-28days postnatally. Aggression was measured in males from both lines, as well as control littermates, using a resident-intruder behavioral test. Consistent with previous reports, male Oxtr-/- mice had elevated levels of aggression relative to controls. Oxtr FB/FB mice on the other hand displayed levels of aggression similar to control animals. In addition, following a resident-intruder test, Oxtr+/+ mice that displayed aggression had less c-fos immunoreactivity in the ventral portion of the lateral septum than those that did not. Further, Oxtr-/- mice had increased c-fos immunoreactivity in the medial amygdala relative to controls. These data suggest that Oxt may play an important role during development in the organization of the neural circuits that underlie aggressive behavior in adulthood, with its absence resulting in heightened aggression.

Exceptional stories of microRNAs

MicroRNAs (miRNAs) are small, endogenous, non-coding RNAs that regulate the expression of target genes in eukaryotic cells and have been extensively studied in the past decade. However, recent evidence suggests that miRNAs have additional, important roles and functions other than post-transcriptional regulation through binding at the 3′ untranslated regions of their target genes. This review describes newly discovered information about the biogenesis and functions of miRNAs as well as presents different points of view about the miRNA system. Our increasing knowledge of the exceptional stories of miRNAs will offer new insights into these powerful gene regulators from virus and bacteria to animals.

MicroRNA profiling in the mouse hypothalamus reveals oxytocin-regulating microRNA.

Oxytocin (Oxt), produced in the hypothalamic paraventricular and supraoptic nuclei for transport to and release from the posterior pituitary, was originally discovered through its role in lactation and parturition. Oxt also plays important roles in the central nervous system by influencing various behaviors. MicroRNAs (miRNAs), endogenous regulators of many genes, are a class of small non‐coding RNAs that mediate post‐transcriptional gene silencing. We performed miRNA expression profiling of the mouse hypothalamus by deep sequencing. Among the sequenced and cross‐mapped small RNAs, expression of known miRNAs and unknown miRNAs candidates were analyzed. We investigated in detail one miRNA, miR‐24, and found that it is a novel regulator of Oxt and controls both transcript and peptide levels of Oxt. These results provide insights into potential neurohypophysial hormone regulation mediated by miRNAs.

Effect of garlic on bacterial biofilm formation on orthodontic wire.

OBJECTIVE To examine the effect of garlic extract on the biofilm formation by Streptococcus mutans on orthodontic wire and on glucosyltransferase gene expression. MATERIALS AND METHODS Growth inhibition of oral bacteria was tested after 50 µL of garlic extract was placed on an agar plate. The minimum inhibitory concentration (MIC) of garlic extract on S mutans growth was first determined. After cultivating streptococci in biofilm medium (BM)-sucrose with garlic extract and orthodontic wire, adenosine triphosphate (ATP) measurement and viable cell counting was performed from the bacteria attached on the wire. Scanning electron microscopy (SEM) analysis of morphology was observed on bacterial cells attached to orthodontic wire. The effect of garlic extract on gene expression was evaluated using quantitative real-time polymerase chain reaction (PCR) of glucosyltransferase. RESULTS Though garlic extract had a clear antibacterial effect on all microorganisms, it also enhanced S mutans attachment on orthodontic wire. Low concentration of garlic extract also increased glucosyltransferase gene expression of S mutans. CONCLUSIONS Despite its antibacterial function, garlic extract increases biofilm formation by S mutans to orthodontic wire, likely through upregulation of glucosyltransferase expression. Garlic extract may thus play an important role in increased bacterial attachment to orthodontic wires.