Effect of Companion Presence during Skin-to-Skin Contact on Maternal Anxiety: A Randomized Clinical Trial.

OBJECTIVE: To compare the effect of companion presence versus midwife presence during skin-to-skin contact (SSC) at birth on maternal anxiety and satisfaction, and neonatal physiological parameters. METHODS: A randomized controlled trial was conducted on 92 pregnant women who were randomized to provide SSC to their newly borns for one hour postpartum, in the presence of a companion (study group) or a midwife (control group). Maternal anxiety (using the Visual Analogue Scale) and the neonatal physiological parameters (including temperature, heart rate, and oxygen saturation) were assessed in four stages viz., immediately after birth, and at 30, 60 and 90 minutes after birth. Maternal satisfaction was also evaluated after transferring the mother to the postpartum ward. RESULTS: We analyzed 86 mother-infant dyads (43 per group). Having a companion significantly reduced maternal anxiety after birth as compared to having a midwife at 30, 60, and 90 minutes after birth (P = 0.04, P = 0.01, and P = 0.04, respectively). There was also a small to medium effect size of the presence of companion compared to midwife in terms of maternal anxiety at 30 minutes (Cohen's d = 0.45; 95% CI = 0.02, 0.87), 60 minutes (Cohen's d = 0.52; 95% CI = 0.08, 0.94) and 90 minutes after birth (Cohen's d = 0.45; 95% CI = 0.02, 0.88). However, there was no significant effect of the same on neonatal physiological parameters. Having a companion versus a midwife led to higher maternal satisfaction rates (P = 0.02); 65.1% of mothers in the study group and 37.2% of mothers in the control group were desirous of the same care in future (P = 0.02). CONCLUSION: Companion presence during SSC leads to a significant reduction in maternal anxiety compared to midwife presence.

A succinate/SUCNR1-brush cell defense program in the tracheal epithelium.

Host-derived succinate accumulates in the airways during bacterial infection. Here, we show that luminal succinate activates murine tracheal brush (tuft) cells through a signaling cascade involving the succinate receptor 1 (SUCNR1), phospholipase Cß2, and the cation channel transient receptor potential channel subfamily M member 5 (TRPM5). Stimulated brush cells then trigger a long-range Ca2+ wave spreading radially over the tracheal epithelium through a sequential signaling process. First, brush cells release acetylcholine, which excites nearby cells via muscarinic acetylcholine receptors. From there, the Ca2+ wave propagates through gap junction signaling, reaching also distant ciliated and secretory cells. These effector cells translate activation into enhanced ciliary activity and Cl- secretion, which are synergistic in boosting mucociliary clearance, the major innate defense mechanism of the airways. Our data establish tracheal brush cells as a central hub in triggering a global epithelial defense program in response to a danger-associated metabolite.

Investigation of the Relationship Between Religious Attitude and Postpartum Physical and Perineal Pain in Iran.

Postpartum pain is one of the most important and common problems of postpartum mothers who resort to non-pharmacological strategies to relieve it due to the side effects of painkillers. This study was conducted to investigate the relationship between religiosity and postpartum physical and perineal pain. The sample size in this cross-sectional study was 101 women giving birth in educational and medical centers of Shiraz, Iran, that was selected by purposive-convenience sampling. Then, a questionnaire was completed for each of them including demographic characteristics, religious attitude, and visual scale of postpartum pain. Data were analyzed by SPSS software. The frequency of the majority of the research population was found at moderate religious attitudes 65.4% (72 individuals). There is a positive and Statistical inverse correlation between religious attitude and perineal pain. According to Fisher's test, in mothers with a high religious attitude, 9.7% had severe perineal pain, 41.9% had moderate pain, and 48.4% had mild pain (p = 0.001). Besides, in the case of physical pain, 3.2%, 35.5%, and 61.3% had severe, moderate, and mild pain, respectively. Regarding physical pains, with the increase in the level of mothers' religious attitude, physical pains also decreased, but the statistical relationship was not significant (p = 0.32). The results showed that the religiosity and spirituality of pregnant women have relieving effects on postpartum pain. Therefore, more attention to the spiritual dimension of human existence and planning to improve it using prayer therapy, dhikr, and meditation is suggested as a strategy to deal with the fear of pain and childbirth and reduce psychological and physical changes before and after childbirth. These findings apply to women with ectopic pregnancies in Iran.

Aging - What it is and how to measure it.

The current understanding of the biology of aging is largely based on research aimed at identifying factors that influence lifespan. However, lifespan as a sole proxy measure of aging has limitations because it can be influenced by specific pathologies (not generalized physiological deterioration in old age). Hence, there is a great need to discuss and design experimental approaches that are well-suited for studies targeting the biology of aging, rather than the biology of specific pathologies that restrict the lifespan of a given species. For this purpose, we here review various perspectives on aging, discuss agreement and disagreement among researchers on the definition of aging, and show that while slightly different aspects are emphasized, a widely accepted feature, shared across many definitions, is that aging is accompanied by phenotypic changes that occur in a population over the course of an average lifespan. We then discuss experimental approaches that are in line with these considerations, including multidimensional analytical frameworks as well as designs that facilitate the proper assessment of intervention effects on aging rate. The proposed framework can guide discovery approaches to aging mechanisms in all key model organisms (e.g., mouse, fish models, D. melanogaster, C. elegans) as well as in humans.

The effect of Pilates exercises on sleep quality and fatigue among female students dormitory residents.

BACKGROUND: This study aims to investigate the effect of Pilates exercises on sleep and fatigue among female college students residing in the dormitory. METHODS: This quasi-experimental study, two parallel groups was performed on 80 single female college students (40 per group), between 18 to 26 years old who lived in the two dormitories. One dormitory was considered as the intervention group and another as the control group. The Pilates group received three one-hour Pilates exercise sessions per week for eight weeks and the control group maintained their routine activities. The Pittsburgh Sleep Quality Index (PSQI) and the Multidimensional Fatigue Inventory (MFI-20) were used respectively to assess sleep quality and fatigue levels, at three time points: baseline, end of week four, and eight follow-ups. Fisher's exact, Chi-square, independent sample t-test and repeated measurements were used. RESULTS: Overall, 66 participants completed the study (32 and 35 participants in the Pilates and control groups, respectively). After four and eight weeks of intervention, the overall mean score of sleep quality improved significantly (p < 0.001). At week four of the intervention, the Pilates group had a significantly lower mean score for subjective sleep quality and daytime dysfunction than the control group (p < 0.001 and p < 0.002, respectively), although sleep duration and habitual sleep efficiency improved after eight weeks of intervention (p < 0.04 and p < 0.034, respectively). Additionally, the overall mean score of fatigue and its dimensions in weeks four and eight of the intervention in the Pilates group were significantly lower compared to the control group (p < 0.001). CONCLUSION: After eight weeks of Pilates exercises implementation, most components of sleep quality significantly improved; however, the effect of Pilates exercises on fatigue was evident from week four onward. Trial registration This trial was registered on 2/6/2015 in the Iranian Registry of Clinical Trials with the IRCT ID: IRCT201412282324N15. URL of registry: https://www.irct.ir/trial/1970 .

The effect of combined knockdowns of Attacins on survival and bacterial load in Tenebrio molitor.

Introduction: Upon infection, insect hosts simultaneously express a cocktail of antimicrobial peptides (AMPs) which can impede pathogen colonization and increase host fitness. It has been proposed that such a cocktail might be adaptive if the effects of co-expressed AMPs are greater than the sum of individual activities. This could potentially prevent the evolution of bacterial resistance. However, in vivo studies on AMPs in combination are scarce. Attacins are one of the relatively large AMP families, which show anti-Gram-negative activity in vitro. Material and methods: Here, we used RNA interference (RNAi) to silence three members of the Attacin family genes in the mealworm beetle, Tenebrio molitor: (TmAttacin1a (TmAtt1a), TmAttacin1b (TmAtt1b), and TmAttacin2 (TmAtt2) both individually and in combination. We then infected T. molitor with the Gram negative entomopathogen Pseudomonas entomophila. Results: We found that survival of the beetles was only affected by the knockdown of TmAttacin1b, TmAttacin2 and the knockdown of all three Attacins together. Triple knockdown, rather than individual or double knockdowns of AMPs, changes the temporal dynamics of their efficiency in controlling the colonization of P. entomophila in the insect body. Discussion: More precisely, AMP gene expression influences P. entomophila load early in the infection process, resulting in differences in host survival. Our results highlight the importance of studying AMP-interactions in vivo.

Abortion services during the COVID-19 pandemic: a systematic review.

Evidence suggests that COVID-19 may impair access to sexual and reproductive health services and safe abortion. The purpose of this systematic review was investigating the changes of abortion services in the COVID-19 pandemic era. We searched PubMed, Web of Science and Scopus for relevant studies published as of August 2021, using relevant keywords. RCT and non-original studies were excluded from the analysis and 17 studies of 151 included in our review. Requests to access medication abortion by telemedicine and demand for self-managed abortion were the main findings of identified studies. Women requested an abortion earlier in their pregnancy, and were satisfied with tele-abortion care due to its flexibility, and ongoing telephone support. Presenting telemedicine services without ultrasound has also been reported. Visits to clinics were reduced based on the severity of the restrictions, and abortion clinics had less revenue, more costs, and more changes in the work style of their healthcare providers. Telemedicine was reported safe, effective, acceptable, and empowering for women. Reasons for using tele-abortion were privacy, secrecy, comfort, using modern contraception, employing of women, distance from clinics, travel restrictions, lockdowns, fear of COVID-19, and political reasons (abortion prohibition). Complications of women using tele-abortion were pain, lack of psychological support, bleeding, and need to blood transfusions. The results of this study showed that using telemedicine and teleconsultations for medical abortion in the pandemic conditions may be extended after pandemic. Findings can be used by reproductive healthcare providers and policy makers to address the complications of abortion services.Trail registration This study is registered in PROSPERO with number CRD42021279042.

COVID-19 pandemic shocks the international community, especially health policymakers around the world. The most important consequence of this outbreak has been direct and indirect impacts on health service provisions in all parts of the health system, including sexual and reproductive health services. We reviewed numerous studies investigating healthcare related to abortion in the pandemic era that showed women had more requests to access medical abortion, more than surgical. They preferred self-managed abortion process by telemedicine. Presenting telemedicine services without ultrasound has also been reported. Visits to clinics were reduced, and this decrease was reported based on the severity of the restrictions. Abortion clinics had reduced revenue, increased costs, and changed work style of their healthcare providers. Reasons for using telemedicine were fear of COVID-19, travel restrictions, lockdowns, more privacy, secrecy, and comfort. Telemedicine was reported safe, effective, acceptable, satisfying, and empowering for women. Maternal complications using tele-abortion were pain, bleeding, and need to blood transfusions. These findings can be used by policy makers and reproductive healthcare providers to address the complications of abortion management.

The relationship between sex steroids (E2, progesterone, and AMH) levels and severity and fatality of COVID-19: A systematic review.

Sex steroids are powerful modulators of the immune system and they may affect the immune response and inflammatory consequences of COVID-19. This systematic review aims to explore the impact of sex steroids on COVID-19 mortality and complications. We looked up the keywords of the study in Scopus, PubMed, and Web of Science. All related original articles published in English, as of October 16, 2021, were reviewed to be included in our research. Concerns regarding the effect of sex hormones on COVID-19, eight full texts have been identified for the conclusion. In these studies, the relationship between estradiol and COVID-19 mortality has been mentioned. The most significant findings were the higher COVID-19 mortality rate in men, compared to women; also, in menopausal women compared to younger women and who received estradiol. In two studies, oral contraceptive pills had a protective effect on the morbidity of SARS-CoV-2 infection. In a randomized controlled trial, subcutaneous injection of progesterone in hospitalized men significantly reduced their symptoms and need for oxygen therapy. Hormone replacement therapy was positively associated with reducing COVID-19 symptoms. Although the results were insufficient for a conclusion, this study represents estrogen as an appropriate pharmacological method for preventing and diminishing the inflammation related to COVID-19 disease. However, future prospective studies and clinical trials are needed to clarify and approve this protective effect.

The effect of vitamin D on the severity of dysmenorrhea and menstrual blood loss: a randomized clinical trial.

BACKGROUND: Primary dysmenorrhea is considered as one of the women's main problems during reproductive age. The present study aimed to investigate the effect of vitamin D on the severity of dysmenorrhea and menstrual blood loss. METHODS: This double-blind, randomized, placebo-controlled trial, was performed on 84 single female college students between 18 and 25 years old who living in dormitories. Students with primary dysmenorrhea and vitamin D deficiency were divided into experimental (n = 42) and control (n = 42) groups. Five days before the putative beginning of their next menstrual cycle, the experimental group received 300,000 IU vitamin D (50,000 IU, two tablets every 8 h), and the control group received a placebo (oral paraffin). The effects of the supplement on the severity of dysmenorrhea and menstrual blood loss were evaluated one cycle before and during two successive cycles. Using the visual analog scale (VAS), verbal multidimensional scoring system (VMS), and pictorial blood assessment chart (PBLAC) questionnaires. Fisher's exact, Chi-square, independent sample t-test and repeated measurements were used. RESULTS: In total, 78 of the 84 students completed the study (39 students per group). The intervention resulted in a significant reduction in the mean scores of both the VAS and VMS in the experimental group, in the first and second menstrual cycles (p < 0.001, p < 0.001, respectively), but not in the means score of PBLAC. Mefenamic acid consumption at the first and second menstruation period, in the experimental group was lower than the control group (p = 0.009, p < 0.001, respectively). CONCLUSIONS: The results indicate that vitamin D supplementation could decrease the severity of primary dysmenorrhea and the need to consume pain-relief medications. Contrariwise vitamin D supplementation had no significant effect on menstrual blood loss. TRIAL REGISTRATION: This trial was registered in the Iranian Registry of Clinical Trials with code IRCT201305212324N on 18/1/2014. URL of registry: https://en.irct.ir/trial/1964 .

Targeting the "hallmarks of aging" to slow aging and treat age-related disease: fact or fiction?

Aging is a major risk factor for a number of chronic diseases, including neurodegenerative and cerebrovascular disorders. Aging processes have therefore been discussed as potential targets for the development of novel and broadly effective preventatives or therapeutics for age-related diseases, including those affecting the brain. Mechanisms thought to contribute to aging have been summarized under the term the "hallmarks of aging" and include a loss of proteostasis, mitochondrial dysfunction, altered nutrient sensing, telomere attrition, genomic instability, cellular senescence, stem cell exhaustion, epigenetic alterations and altered intercellular communication. We here examine key claims about the "hallmarks of aging". Our analysis reveals important weaknesses that preclude strong and definitive conclusions concerning a possible role of these processes in shaping organismal aging rate. Significant ambiguity arises from the overreliance on lifespan as a proxy marker for aging, the use of models with unclear relevance for organismal aging, and the use of study designs that do not allow to properly estimate intervention effects on aging rate. We also discuss future research directions that should be taken to clarify if and to what extent putative aging regulators do in fact interact with aging. These include multidimensional analytical frameworks as well as designs that facilitate the proper assessment of intervention effects on aging rate.

Tachykinin-related peptides modulate immune-gene expression in the mealworm beetle Tenebrio molitor L.

Tachykinins (TKs) are a group of conserved neuropeptides. In insects, tachykinin-related peptides (TRPs) are important modulators of several functions such as nociception and lipid metabolism. Recently, it has become clear that TRPs also play a role in regulating the insect immune system. Here, we report a transcriptomic analysis of changes in the expression levels of immune-related genes in the storage pest Tenebrio molitor after treatment with Tenmo-TRP-7. We tested two concentrations (10-8 and 10-6 M) at two time points, 6 and 24 h post-injection. We found significant changes in the transcript levels of a wide spectrum of immune-related genes. Some changes were observed 6 h after the injection of Tenmo-TRP-7, especially in relation to its putative anti-apoptotic action. Interestingly, 24 h after the injection of 10-8 M Tenmo-TRP-7, most changes were related to the regulation of the cellular response. Applying 10-6 M Tenmo-TRP-7 resulted in the downregulation of genes associated with humoral responses. Injecting Tenmo-TRP-7 did not affect beetle survival but led to a reduction in haemolymph lysozyme-like antibacterial activity, consistent with the transcriptomic data. The results confirmed the immunomodulatory role of TRP and shed new light on the functional homology between TRPs and TKs.

Cysteinyl leukotrienes and acetylcholine are biliary tuft cell cotransmitters.

The gallbladder stores bile between meals and empties into the duodenum upon demand and is thereby exposed to the intestinal microbiome. This exposure raises the need for antimicrobial factors, among them, mucins produced by cholangiocytes, the dominant epithelial cell type in the gallbladder. The role of the much less frequent biliary tuft cells is still unknown. We here show that propionate, a major metabolite of intestinal bacteria, activates tuft cells via the short-chain free fatty acid receptor 2 and downstream signaling involving the cation channel transient receptor potential cation channel subfamily M member 5. This results in corelease of acetylcholine and cysteinyl leukotrienes from tuft cells and evokes synergistic paracrine effects upon the epithelium and the gallbladder smooth muscle, respectively. Acetylcholine triggers mucin release from cholangiocytes, an epithelial defense mechanism, through the muscarinic acetylcholine receptor M3. Cysteinyl leukotrienes cause gallbladder contraction through their cognate receptor CysLTR1, prompting emptying and closing. Our results establish gallbladder tuft cells as sensors of the microbial metabolite propionate, initiating dichotomous innate defense mechanisms through simultaneous release of acetylcholine and cysteinyl leukotrienes.

Tenebrio molitor Spätzle 1b Is Required to Confer Antibacterial Defense Against Gram-Negative Bacteria by Regulation of Antimicrobial Peptides.

Innate immunity is the ultimate line of defense against invading pathogens in insects. Unlike in the mammalian model, in the insect model, invading pathogens are recognized by extracellular receptors, which activate the Toll signaling pathway through an extracellular serine protease cascade. In the Toll-NF-κB pathway, the extracellular spätzle protein acts as a downstream ligand for Toll receptors in insects. In this study, we identified a novel Spätzle isoform (TmSpz1b) from RNA sequencing database of Tenebrio molitor. TmSpz1b was bioinformatically analyzed, and functionally characterized for the antimicrobial function by RNA interference (RNAi). The 702 bp open reading frame of TmSpz1b encoded a putative protein of 233 amino acid residues. A conserved cystine-knot domain with seven cysteine residues in TmSpz1b was involved in three disulfide bridges and the formation of a spätzle dimer. TmSpz1b was mostly expressed in the hemocytes of T. molitor late instar larvae. The mRNA expression of TmSpz1b was highly induced in the hemocytes after Escherichia coli, Staphylococcus aureus, and Candida albicans stimulation of T. molitor larvae. TmSpz1b silenced larvae were significantly more susceptible to E. coli infection. In addition, RNAi-based functional assay characterized TmSpz1b to be involved in the positive regulation of antimicrobial peptide genes in hemocytes and fat bodies. Further, the TmDorX2 transcripts were downregulated in TmSpz1b silenced individuals upon E. coli challenge suggesting the relationship to Toll signaling pathway. These results indicate that TmSpz1b is involved in the T. molitor innate immunity, causes the sequestration of Gram-negative bacteria by the regulatory action of antimicrobial peptides, and enhances the survival of T. molitor larvae.

Novel implications of a strictly monomorphic (GCC) repeat in the human PRKACB gene.

PRKACB (Protein Kinase CAMP-Activated Catalytic Subunit Beta) is predominantly expressed in the brain, and regulation of this gene links to neuroprotective effects against tau and Aß-induced toxicity. Here we studied a (GCC)-repeat spanning the core promoter and 5' UTR of this gene in 300 human subjects, consisting of late-onset neurocognitive disorder (NCD) (N = 150) and controls (N = 150). We also implemented several models to study the impact of this repeat on the three-dimensional (3D) structure of DNA. While the PRKACB (GCC)-repeat was strictly monomorphic at 7-repeats, we detected two 7/8 genotypes only in the NCD group. In all examined models, the (GCC)7 and its periodicals had the least range of divergence variation on the 3D structure of DNA in comparison to the 8-repeat periodicals and several hypothetical repeat lengths. A similar inert effect on the 3D structure was not detected in other classes of short tandem repeats (STRs) such as GA and CA repeats. In conclusion, we report monomorphism of a long (GCC)-repeat in the PRKACB gene in human, its inert effect on DNA structure, and enriched divergence in late-onset NCD. This is the first indication of natural selection for a monomorphic (GCC)-repeat, which probably evolved to function as an "epigenetic knob", without changing the regional DNA structure.

Natural copy number variation of tandemly repeated regulatory SNORD RNAs leads to individual phenotypic differences in mice.

Genic copy number differences can have phenotypic consequences, but so far this has not been studied in detail in natural populations. Here, we analysed the natural variation of two families of tandemly repeated regulatory small nucleolar RNAs (SNORD115 and SNORD116) in the house mouse (Mus musculus). They are encoded within the Prader-Willi Syndrome gene region, known to be involved in behavioural, metabolic, and osteogenic functions in mammals. We determined that the copy numbers of these SNORD RNAs show substantial natural variation, both in wild-derived mice as well as in an inbred mouse strain (C57BL/6J). We show that copy number differences are subject to change across generations, making them highly variable and resulting in individual differences. In transcriptome data from brain samples, we found SNORD copy-number correlated regulation of possible target genes, including Htr2c, a predicted target gene of SNORD115, as well as Ankrd11, a predicted target gene of SNORD116. Ankrd11 is a chromatin regulator, which has previously been implicated in regulating the development of the skull. Based on morphometric shape analysis of the skulls of individual mice of the inbred strain, we show that shape measures correlate with SNORD116 copy numbers in the respective individuals. Our results suggest that the variable dosage of regulatory RNAs can lead to phenotypic variation between individuals that would typically have been ascribed to environmentally induced variation, while it is actually encoded in individual differences of copy numbers of regulatory molecules.

Autophagy in Tenebrio molitor Immunity: Conserved Antimicrobial Functions in Insect Defenses.

The yellow mealworm beetle (Tenebrio molitor) has been exploited as an experimental model to unravel the intricacies of cellular and humoral immunity against pathogenic infections. Studies on this insect model have provided valuable insights into the phenotypic plasticity of immune defenses against parasites and pathogens. It has thus been possible to characterize the hemocoelic defenses of T. molitor that rely on the recognition of non-self-components of pathogens by pattern recognition receptors (PRRs). The subsequent signaling cascade activating pathways such as the NF-κB controlled by Toll and IMD pathways lead to the synthesis of antimicrobial peptides (AMPs), onset of hemocyte-driven phagocytosis, and activation of the prophenoloxidase cascade regulating the process of melanization. Nevertheless, the activation of autophagy-mediated defenses of T. molitor against the facultative intracellular gram-positive bacterium Listeria monocytogenes provides clear evidence of the existence of a cross-talk between autophagy and the IMD pathway. Moreover, the identification of several autophagy-related genes (Atgs) in T. molitor transcriptome and expressed sequence tag (EST) databases has contributed to the understanding of the autophagy-signaling cascade triggered by L. monocytogenes challenge. Providing further evidence of the cross-talk hypothesis, TmRelish has been shown to be required not only for regulating the synthesis of AMPs through the PGRP-LE/IMD pathway activation but also for the expression of Atgs in T. molitor larvae following L. monocytogenes challenge. Notably, L. monocytogenes can stimulate the T. molitor innate immune system by producing molecules recognized by the multifunctional PRR (TmPGRP-LE), which stimulates intracellular activation of the IMD pathway and autophagy. Considering the conservation of autophagy components involved in combating intracellular pathogens, it will be interesting to extrapolate a dynamic cross-talk model of immune activation. This review summarizes the most significant findings on the regulation of autophagy in T. molitor during L. monocytogenes infection and on the role of the innate immunity machinery, including the NF-κB pathway, in the control of pathogenic load.

The imprinted lncRNA Peg13 regulates sexual preference and the sex-specific brain transcriptome in mice.

Mammalian genomes include many maternally and paternally imprinted genes. Most of these are also expressed in the brain, and several have been implicated in regulating specific behavioral traits. Here, we have used a knockout approach to study the function of Peg13, a gene that codes for a fast-evolving lncRNA (long noncoding RNA) and is part of a complex of imprinted genes on chromosome 15 in mice and chromosome 8 in humans. Mice lacking the 3' half of the transcript look morphologically wild-type but show distinct behavioral differences. They lose interest in the opposite sex, instead displaying a preference for wild-type animals of the same sex. Further, they show a higher level of anxiety, lowered activity and curiosity, and a deficiency in pup retrieval behavior. Brain RNA expression analysis reveals that genes involved in the serotonergic system, formation of glutamatergic synapses, olfactory processing, and estrogen signaling-as well as more than half of the other known imprinted genes-show significant expression changes in Peg13-deficient mice. Intriguingly, these pathways are differentially affected in the sexes, resulting in male and female brains of Peg13-deficient mice differing more from each other than those of wild-type mice. We conclude that Peg13 is part of a developmental pathway that regulates the neurobiology of social and sexual interactions.

Critical Roles of Spätzle5 in Antimicrobial Peptide Production Against Escherichia coli in Tenebrio molitor Malpighian Tubules.

The dimeric cytokine ligand Spätzle (Spz) is responsible for Toll pathway activation and antimicrobial peptide (AMP) production upon pathogen challenge in Tenebrio molitor. Here, we indicated that TmSpz5 has a functional role in response to bacterial infections. We showed that the highest expression of TmSpz5 is induced by Candida albicans. However, TmSpz5 knockdown reduced larval survival against Escherichia coli and Staphylococcus aureus. To evaluate the molecular mechanism underlying the observed survival differences, the role of TmSpz5 in AMP production was examined by RNA interference and microbial injection. T. molitor AMPs that are active against Gram-negative and -positive bacteria, including Tmtenecins, Tmattacins, Tmcoleoptericins, Tmtaumatin-like-proteins, and Tmcecropin-2, were significantly downregulated by TmSpz-5 RNAi in the Malpighian tubules (MTs) following a challenge with E. coli and S. aureus. However, upon infection with C. albicans the mRNA levels of most AMPs in the dsTmSpz5-injected group were similar to those in the control groups. Likewise, the expression of the transcription factors NF-κB, TmDorX2, and TmRelish were noticeably suppressed in the MTs of TmSpz5-silenced larvae. Moreover, E. coli-infected TmSpz5 knockdown larvae showed decreased antimicrobial activity in the MTs and hindgut compared with the control group. These results demonstrate that TmSpz5 has a defined role in T. molitor innate immunity by regulating AMP expression in MTs in response to E. coli.

Evolution of a New Testis-Specific Functional Promoter Within the Highly Conserved Map2k7 Gene of the Mouse.

Map2k7 (synonym Mkk7) is a conserved regulatory kinase gene and a central component of the JNK signaling cascade with key functions during cellular differentiation. It shows complex transcription patterns, and different transcript isoforms are known in the mouse (Mus musculus). We have previously identified a newly evolved testis-specific transcript for the Map2k7 gene in the subspecies M. m. domesticus. Here, we identify the new promoter that drives this transcript and find that it codes for an open reading frame (ORF) of 50 amino acids. The new promoter was gained in the stem lineage of closely related mouse species but was secondarily lost in the subspecies M. m. musculus and M. m. castaneus. A single mutation can be correlated with its transcriptional activity in M. m. domesticus, and cell culture assays demonstrate the capability of this mutation to drive expression. A mouse knockout line in which the promoter region of the new transcript is deleted reveals a functional contribution of the newly evolved promoter to sperm motility and the spermatid transcriptome. Our data show that a new functional transcript (and possibly protein) can evolve within an otherwise highly conserved gene, supporting the notion of regulatory changes contributing to the emergence of evolutionary novelties.

Dedicated transcriptomics combined with power analysis lead to functional understanding of genes with weak phenotypic changes in knockout lines.

Systematic knockout studies in mice have shown that a large fraction of the gene replacements show no lethal or other overt phenotypes. This has led to the development of more refined analysis schemes, including physiological, behavioral, developmental and cytological tests. However, transcriptomic analyses have not yet been systematically evaluated for non-lethal knockouts. We conducted a power analysis to determine the experimental conditions under which even small changes in transcript levels can be reliably traced. We have applied this to two gene disruption lines of genes for which no function was known so far. Dedicated phenotyping tests informed by the tissues and stages of highest expression of the two genes show small effects on the tested phenotypes. For the transcriptome analysis of these stages and tissues, we used a prior power analysis to determine the number of biological replicates and the sequencing depth. We find that under these conditions, the knockouts have a significant impact on the transcriptional networks, with thousands of genes showing small transcriptional changes. GO analysis suggests that A930004D18Rik is involved in developmental processes through contributing to protein complexes, and A830005F24Rik in extracellular matrix functions. Subsampling analysis of the data reveals that the increase in the number of biological replicates was more important that increasing the sequencing depth to arrive at these results. Hence, our proof-of-principle experiment suggests that transcriptomic analysis is indeed an option to study gene functions of genes with weak or no traceable phenotypic effects and it provides the boundary conditions under which this is possible.