A Farnesyltransferase Inhibitor Restores Cognitive Deficits in Tsc2+/- Mice through Inhibition of Rheb1.

Tuberous sclerosis complex (TSC) is caused by mutations in Tsc1 or Tsc2, whose gene products inhibit the small G-protein Rheb1. Rheb1 activates mTORC1, which may cause refractory epilepsy, intellectual disability, and autism. The mTORC1 inhibitors have been used for TSC patients with intractable epilepsy. However, its effectiveness for cognitive symptoms remains unclear. We found a new signaling pathway for synapse formation through Rheb1 activation, but not mTORC1. Here, we show that treatment with the farnesyltransferase inhibitor lonafarnib increased unfarnesylated (inactive) Rheb1 levels and restored synaptic abnormalities in cultured Tsc2+/- neurons, whereas rapamycin did not enhance spine synapse formation. Lonafarnib treatment also restored the plasticity-related Arc (activity-regulated cytoskeleton-associated protein) expression in cultured Tsc2+/- neurons. Lonafarnib action was partly dependent on the Rheb1 reduction with syntenin. Oral administration of lonafarnib increased unfarnesylated protein levels without affecting mTORC1 and MAP (mitogen-activated protein (MAP)) kinase signaling, and restored dendritic spine morphology in the hippocampi of male Tsc2+/- mice. In addition, lonafarnib treatment ameliorated contextual memory impairments and restored memory-related Arc expression in male Tsc2+/- mice in vivo Heterozygous Rheb1 knockout in male Tsc2+/- mice reproduced the results observed with pharmacological treatment. These results suggest that the Rheb1 activation may be responsible for synaptic abnormalities and memory impairments in Tsc2+/- mice, and its inhibition by lonafarnib could provide insight into potential treatment options for TSC-associated neuropsychiatric disorders.SIGNIFICANCE STATEMENT Tuberous sclerosis complex (TSC) is an autosomal-dominant disease that causes neuropsychiatric symptoms, including intractable epilepsy, intellectual disability (ID) and autism. No pharmacological treatment for ID has been reported so far. To develop a pharmacological treatment for ID, we investigated the mechanism of TSC and found that Rheb1 activation is responsible for synaptic abnormalities in TSC neurons. To inhibit Rheb1 function, we used the farnesyltransferase inhibitor lonafarnib, because farnesylation of Rheb1 is required for its activation. Lonafarnib treatment increased inactive Rheb1 and recovered proper synapse formation and plasticity-related Arc (activity-regulated cytoskeleton-associated protein) expression in TSC neurons. Furthermore, in vivo lonafarnib treatment restored contextual memory and Arc induction in TSC mice. Together, Rheb1 inhibition by lonafarnib could provide insight into potential treatments for TSC-associated ID.

The preventive effects of perioperative oral care on surgical site infections after pancreatic cancer surgery: a retrospective study.

PURPOSE: Pancreatic ductal adenocarcinoma (PDAC) is the most malignant cancer of the gastrointestinal system, and is associated with high rates of postoperative complications, including surgical site infections (SSIs). Perioperative oral care is an effective measure for preventing postoperative pneumonia. However, the preventive effects of perioperative oral care on SSIs have not been reported. We investigated the preventive effects of perioperative oral care on SSIs after pancreatic cancer surgery. METHODS: A total of 103 patients with PDAC who underwent radical resection at Hiroshima Prefectural Hospital (2011-2018) were enrolled in this retrospective study. Of the 103 patients, 75 received perioperative oral care by dentists and dental hygienists (oral care group), whereas 28 did not (control group). Univariate and multivariate analyses with propensity score as a covariate were used to investigate the incidence and risk factors of SSIs in the oral care and control groups. RESULTS: The incidence of SSIs was significantly lower in the oral care group than in the control group (12.0% vs. 39.3%, P = 0.004). Logistic regression analysis revealed that a soft pancreas, the surgical procedure (pancreaticoduodenectomy), blood transfusion, diabetes mellitus, and the absence of oral care intervention were risk factors for SSIs. The odds ratio for the absence of oral care intervention was 6.090 (95% confidence interval: 1.750-21.200, P = 0.004). CONCLUSION: Our results suggest that perioperative oral care may reduce the risk of developing SSIs after pancreatic cancer surgery. These findings need to be evaluated in future prospective studies. TRIAL REGISTRATION: UMIN registration number: UMIN000042082; October 15, 2020, retrospectively registered.

Secondary school students' and peer educators' perceptions of adolescent education in rural Tanzania: a qualitative study.

BACKGROUND: In many African countries, cultural norms and values hinder conversations about sexuality among adolescents and their parents. Currently, there are no sex education classes in the curriculum at schools in Tanzania. Even when sex education is provided, the content is often abstinence-oriented, and there is a lack of in-depth instruction and exploration on the topic. To help overcome this, peer education is encouraged. After implementing peer-based adolescent education via a non-profit organization, this study aims to (1) identify students' and peer educators' perceptions of adolescent education and (2) identify the changes that occur as a result of adolescent education with peer educators. METHODS: This was a qualitative descriptive study using focus group discussions (FGDs). Secondary school students, including peer educators as well as students who received adolescent education, were asked about their perception of peer-based adolescent education. The FGDs were conducted in Swahili with the support of local collaborators. Data were transcribed and translated into English and Japanese. Content analysis was conducted to merge the categories and subcategories. RESULTS: A total of 92 students (57 girls and 35 boys) were included from three urban and three rural secondary schools where peer education was being implemented. Six FGDs were conducted for girls and four for boys, for a total of 10 FGDs. The students had both positive and negative perceptions of peer-based adolescent education. Both the peer educators and the other students felt that they gained more confidence through the process, based on the conversations they had and the trusting relationship that formed as a result. The peer educators were also successful in eliciting behavioral changes, and the students shared their sex-related knowledge with other peers as well. CONCLUSION: The peer education process helped students gain confidence in teaching their peers and elicit behavioral changes. Adult supervision for peer educators is suggested.

Peer education, such as sharing correct knowledge, skills, and behaviors, is encouraged for maintaining a healthy lifestyle in later life. The non-profit organization Class for Everyone conducted adolescent education in collaboration with a local NGO, the New Rural Children Foundation, to prevent unwanted pregnancy and social isolation among adolescent girls. In secondary schools, the NGO members provided adolescent education led by peer educators.In this study, we conducted focus group discussions to understand how peer educators and other secondary school students perceived peer-based adolescent education. The study included 92 students (57 girls and 35 boys) from three urban and three rural secondary schools where peer education was being implemented. We used content analysis to merge the categories and subcategories.We found that students had both positive and negative perceptions about peer-based adolescent education. The participants felt that they gained more confidence through the peer education process. The peer educators were also successful in eliciting behavioral changes. Moreover, the students receiving peer education shared their sex-related knowledge with other peers as well.In conclusion, the peer education process helped students gain confidence in teaching their peers and elicit behavioral change. Adult supervision for peer educators is suggested.

Birth preparedness and related factors: a cross-sectional study in Tanzania City area.

BACKGROUND: Birth preparedness could be the key factor that influences the choice of birthplace with skilled birth attendants. To reduce the high maternal mortality of Tanzania, a large study was planned to develop a smartphone app to promote birth preparedness in a city area of Tanzania. This study aimed to identify factors that influence birth preparedness in the city area of Tanzania. METHODS: Pregnant women were asked to complete the Birth Preparedness Questionnaire during antenatal visits using tablets. Multiple linear regression analyses were performed to determine the sociodemographic and obstetric characteristics that influenced the factors. RESULTS: A total of 211 participants were included in the analysis. Distance from the nearest health facility negatively influenced the total score of the Birth Preparedness Assessment (ß= 0.7, p = 0.02). Education higher than college positively influenced the total score (ß = 4.76, p = 0.01). Decision-making of birthplace by other people (not women) negatively influenced Family Support (ß=1.18, p = 0.03). Having jobs negatively influenced Preparation of Money and Food (ß=-1.02, p < 0.01) and positively influenced the knowledge (ß = 0.75, p = 0.03). Being single positively influenced Preparation of Money and Food (ß = 0.35, p = 0.19) and Preference of Skilled Birth Attendants (ß = 0.42, p = 0.04). Experience of losing a baby negatively influenced the knowledge (ß=0.80, p < 0.01) and Preference of Skilled Birth Attendants (ß=0.38, p = 0.02). CONCLUSIONS: The findings showed an updated information on pregnant Tanzanian women living in an urban area where rapid environmental development was observed. Birth preparedness was negatively affected when women reside far from the health facilities, the birthplace decision-making was taken by others beside the women, women have jobs, and when women have experienced the loss of a baby. We hope to use the information from this study as content in our future study, in which we will be applying a smartphone app intervention for healthy pregnancy and birth preparedness. This information will also help in guiding the analysis of this future study. Although generalization of the study needs careful consideration, it is important to reconsider issues surrounding birth preparedness as women's roles both in the family and society, are more, especially in urban settings.

Primary and immortalized cell lines derived from the Amami rabbit (Pentalagus furnessi) and evolutionally conserved cell cycle control with CDK4 and Cyclin D1.

The Amami rabbit (Pentagulus furnessi) is a dark brown-furred rabbit classified as an endangered species and only found in the Amami Islands of Japan. They are often called living fossils because they retain primitive characteristics of ancient rabbits that lived approximately 1 million years ago, such as short feet and hind legs and small ears. Although the ancient rabbit has disappeared due to the competition with European rabbit (Oryctolagus cuniculus) in the most of the Asian area, Amami rabbit survived since Amami Islands has isolated from Japan and Taiwan. Although Amari rabbit is one of the protected animals, their population decreases each year due to human activities, such as deforestation and roadkill. In this study, we collected roadkill samples of Amami rabbits and established primary and immortalized fibroblast cell lines. Combined expression of human-derived mutant Cyclin-dependent kinase 4, Cyclin D1, and hTERT allowed us to immortalize fibroblasts successfully in three individuals of Amami rabbits. The immortalized fibroblasts dramatically extended the cell culture period, when it was compared with the cell culture period of wild type cells. Furthermore, the immortalized cells maintained their normal chromosomal pattern (2n = 46). Our results suggest that cellular senescence which mainly regulated by p16-RB signaling pathway is conserved in animal evolution at least from 1 million years ago.

A Single Pheromone Receptor Gene Conserved across 400 My of Vertebrate Evolution.

Pheromones are crucial for eliciting social and sexual behaviors in diverse animal species. The vomeronasal receptor type-1 (V1R) genes, encoding members of a pheromone receptor family, are highly variable in number and repertoire among mammals due to extensive gene gain and loss. Here, we report a novel pheromone receptor gene belonging to the V1R family, named ancient V1R (ancV1R), which is shared among most Osteichthyes (bony vertebrates) from the basal lineage of ray-finned fishes to mammals. Phylogenetic and syntenic analyses of ancV1R using 115 vertebrate genomes revealed that it represents an orthologous gene conserved for >400 My of vertebrate evolution. Interestingly, the loss of ancV1R in some tetrapods is coincident with the degeneration of the vomeronasal organ in higher primates, cetaceans, and some reptiles including birds and crocodilians. In addition, ancV1R is expressed in most mature vomeronasal sensory neurons in contrast with canonical V1Rs, which are sparsely expressed in a manner that is consistent with the "one neuron-one receptor" rule. Our results imply that a previously undescribed V1R gene inherited from an ancient Silurian ancestor may have played an important functional role in the evolution of vertebrate vomeronasal organ.

Multiplex Real-Time Loop-Mediated Isothermal Amplification Using an Electrochemical DNA Chip Consisting of a Single Liquid-Flow Channel.

We developed a multiplex system capable of simultaneously quantifying different target sequences by applying an electrochemical DNA chip that consists of single liquid-flow channel with primers designed for loop-mediated isothermal amplification (LAMP). We applied this system for detecting mature microRNAs (miRNAs). miRNAs extracted from serum were enzymatically lengthened to about 100 base pairs by reverse-transcription and elongation reactions. The LAMP primers for amplifying the lengthened miRNAs were adsorbed and immobilized on the surface of the liquid-flow channel at five different positions. A LAMP solution containing the lengthened miRNAs, Tin DNA polymerase, and ruthenium hexaamine (RuHex) as a redox compound was injected into the DNA chip. The electrochemical reaction of RuHex in the LAMP solution was then measured continuously via linear-sweep voltammetry at 65 °C. The LAMP reaction of the positive control revealed that the cathodic peak current of RuHex increased. Additionally, the initial number of miRNA copies was correlated with the time when the cathodic current began to increase. Five miRNAs were simultaneously detected at 103-106 copies per 50 µL within 2 h. We expect these results will be useful for developing a simple and stable electrochemical-based method for the real-time monitoring of miRNAs, while also facilitating the implementation of electrochemical DNA chips for molecular analyses.

Refractory Oral Ulcer Responsive to Infliximab: A Case Report.

PURPOSE: Some oral ulcers are refractory to treatment. Recently, anti-tumor necrosis factor-alpha (anti-TNF-α) agents have been shown to induce relatively long remissions of refractory oral ulcers. METHODS: A 69-year man presented with a sore throat caused by a pharyngeal ulcer in the soft palate and subsequently repeatedly developed oropharyngolaryngeal ulcers and an ulcer in the ileocecal region. Intestinal Behçet disease was suspected, but he did not meet the diagnostic criteria for this condition. Seven months after the initial presentation, he developed a lingual ulcer that did not resolve despite treatment with prednisolone. Given the suspicion of intestinal Behçet disease, 300 mg of infliximab was administered intravenously. RESULTS: After the third dose of infliximab with prednisolone, the ulcer began to improve. He received infliximab every 8 weeks thereafter and developed no new oropharyngolaryngeal ulcers during the 17 months after the first infusion of infliximab. CONCLUSIONS: When a patient has refractory oral ulcers, attention must be paid to digestive symptoms and whether the patient has a systemic disease such as Behçet disease investigated. Furthermore, administration of infliximab to patients with refractory ulcers can be considered even when they do not fulfill the clinical criteria for a systemic disease.

A novel voltammetric approach for real-time electrochemical detection of targeted nucleic acid sequences using LAMP.

We have developed a novel voltammetric DNA chip for real-time electrochemical detection of targeted nucleic acid sequences using loop-mediated isothermal amplification (LAMP) and ruthenium hexaamine (RuHex) as the intercalative redox compound. A GspSSD DNA polymerase was used for LAMP owing to its tolerance of the intercalative redox compound. The electrochemical reaction of 1 mM RuHex in the LAMP solution was measured continuously by linear sweep voltammetry at 65 °C using an electrochemical DNA chip. According to the LAMP reaction of the positive sample, the cathodic peak current of RuHex increased and the cathodic peak potential of RuHex shifted to negative voltage. The initial number of copies of the targeted nucleic acid was correlated with both the time when the cathodic current began to increase and the time when the cathodic potential began to shift rapidly. 103 to 106 copies/50 µL of the targeted nucleic acid were detected quantitatively and the detection limit was 101 copies within an hour. We expect these results to lead to the realization of simple and stable electrochemical real-time monitoring of targeted nucleic acids while also facilitating the implementation of electrochemical DNA chips in molecular testing.

Distribution pattern of the celiac, cranial mesenteric, and caudal mesenteric arteries to the gastrointestinal tract in the common marmoset (Callithrix jacchus).

Gastrointestinal diseases pose problems to captive common marmosets. Therefore, knowledge of the anatomy of the arterial supply to the gastrointestinal tract is an important prerequisite for implementing appropriate veterinary care. The common marmoset's intestinal tract has a well-developed cecum specialized for the fermentative digestion of tree gums. This specialized gastrointestinal tract may have a unique pattern of arterial distribution. This study aimed to elucidate the species-specific gastrointestinal tract arterial anatomy of the common marmoset. We traced the celiac, cranial mesenteric, and caudal mesenteric arteries in six male and nine female common marmosets using the latex injection method. We found that the celiac artery gave rise to the left gastric, common hepatic, splenic, and caudal pancreaticoduodenal arteries. In addition to these arteries, the celiac artery gave origin to the middle colic or jejunal arteries in seven or one cases, respectively. The branches of the cranial mesenteric artery consisted of 3-6 arteries, including the middle colic, caudal pancreaticoduodenal, jejunal, right colic, ileocolic, and ileal arteries, as well as a common trunk of the ventral cecal and ileal branches, and the dorsal cecal and colic branches. In four cases, the cranial mesenteric artery gave rise to the jejunal, ileocolic, and ileal arteries. In one of the 13 cases, the celiac and cranial mesenteric arteries formed a common trunk. The caudal mesenteric artery branched into the left colic, sigmoid, and cranial rectal arteries in all the cases. These findings provide an anatomical basis for gastrointestinal veterinary care of common marmosets.

Anxious about rejection, avoidant of neglect: Infant marmosets tune their attachment based on individual caregiver's parenting style.

Children's secure attachment with their primary caregivers is crucial for physical, cognitive, and emotional maturation. Yet, the causal links between specific parenting behaviors and infant attachment patterns are not fully understood. Here we report infant attachment in New World monkeys common marmosets, characterized by shared infant care among parents and older siblings and complex vocal communications. By integrating natural variations in parenting styles and subsecond-scale microanalyses of dyadic vocal and physical interactions, we demonstrate that marmoset infants signal their needs through context-dependent call use and selective approaches toward familiar caregivers. The infant attachment behaviors are tuned to each caregiver's parenting style; infants use negative calls when carried by rejecting caregivers and selectively avoid neglectful and rejecting caregivers. Family-deprived infants fail to develop such adaptive uses of attachment behaviors. With these similarities with humans, marmosets offer a promising model for investigating the biological mechanisms of attachment security.

Development and pilot test of a smartphone app for midwifery care in Tanzania: A comparative cross-sectional study.

To address Tanzania's high maternal mortality ratio, it is crucial to increase women's access to healthcare. To improve access, the quality of antenatal care needs to be improved. Therefore, we conducted a pilot study of a smartphone app for midwives and examined its potential effects on the learning outcomes of midwives and birth preparedness of pregnant women in Tanzania. This mixed-methods, pilot study provided an educational app for midwives in the intervention group, obtained data about the continuous use of the app, measured midwives' learning outcomes, directed focus group discussions on the usability of the app, and conducted surveys among pregnant women about birth preparedness in the intervention and control groups to evaluate if midwives provided proper information to them. The control group received regular antenatal care and answered the same survey. Participants were 23 midwives who participated in the testing and provided learning outcome data. Twenty-one participated in focus group discussions. Results showed that 87.5% of midwives continued to study with the app two months post-intervention. A mini-quiz conducted after using the app showed a significant increase in mean scores (6.9 and 8.4 points, respectively) and a non-significant increase on the questionnaire on women-centered care (98.6 and 102.2 points, respectively). In the focus group discussions, all midwives expressed satisfaction with the app for several reasons, including comprehensive content, feelings of confidence, and reciprocal communication. There were 207 pregnant women included in the analysis. The intervention group had significantly higher knowledge scores and home-based value scores than did controls. The total scores and other subscales did not show statistical significance for group differences. The results indicate the potential impact of the midwifery education app when it is implemented on a larger scale, especially considering that the results show a potential effect on midwives' learning outcomes.

Infective endocarditis with systemic bacterial embolism caused by Staphylococcus aureus in a free-ranging Amami rabbit (Pentalagus furnessi).

The Amami rabbit (Pentalagus furnessi) is found only on the two islands of Amami-Oshima and Tokunoshima in southwest Japan. It has a primitive appearance and ecology, is an evolutionarily valuable animal and has been assigned to the International Union for Conservation of Nature Red List of Threatened Species. We describe a case with mild purulent wounds on the distal digital skin of both forelimbs and multiple nodular lesions in various organs, including the heart and kidney. Microscopically, the heart lesions were characterized by disruption of the mitral valve and multifocal myocardial necrosis and abscesses due to infection with gram-positive cocci. Similar bacterial infarctions were also found in other organs, including the kidneys. The bacteria were identified as Staphylococcus aureus by immunohistochemical and molecular biological examinations. This first report of infective endocarditis and systemic infarctions caused by S. aureus in an Amami rabbit indicates the importance of monitoring purulent injuries, even if mild, to prevent secondary infections in this species.

Immortalization of primary cells derived from the endangered Ryukyu long-furred rat.

The Ryukyu long-furred rat is an endangered species confined to the southernmost three small islands of Japan (Amami-Oshima, Tokunoshima, and Okinawa). Its population is rapidly decreasing because of roadkill, deforestation, and feral animals. To date, its genomic and biological information are poorly understood. In this study, we successfully immortalized Ryukyu long-furred rat cells by expressing a combination of cell cycle regulators, mutant cyclin-dependent kinase 4 (CDK4R24C) and cyclin D1, together with telomerase reverse transcriptase or an oncogenic protein, the Simian Virus large T antigen. The cell cycle distribution, telomerase enzymatic activity, and karyotype of these two immortalized cell lines were analyzed. The karyotype of the former cell line immortalized with cell cycle regulators and telomerase reverse transcriptase retained the nature of the primary cells, while that of the latter cell line immortalized with the Simian Virus large T antigen had many aberrant chromosomes. These immortalized cells would be valuable for studying the genomics and biology of Ryukyu long-furred rats.

Social interaction influences blood cortisol values and brain aromatase genes in the protandrous false clown anemonefish, Amphiprion ocellaris.

Anemonefish, Amphiprion spp., are socially controlled, protandrous sex changers with a monogamous mating system. Under certain conditions, sexually immature anemonefish with ambisexual gonads differentiate directly into males or females. Formation and maintenance of social rank in a group are considered key requirements for the induction of sex change or differentiation. Generally, each animal living in a social group experiences a different level of social stress in accordance with its social rank, and we hypothesize that the stress situation of individual anemonefish influences its sex determination. Groups of three sexually immature anemonefish were placed into each of five experimental tanks and kept for 10 days to allow for social rank formation and behavioral observation. The fish were then euthanized, and blood and brain samples were collected from each fish. The social rank of each individual was distinguishable from day 1 of the experiment. Aggressive behaviors were most frequent and blood Cortisol values were higher in dominant individuals. The transcription of mRNA for stress-related genes, i.e., those encoding for glucocorticoid and arginine vasotocin receptors, was higher in the brains of dominant individuals than in other social ranks. Furthermore, we detected higher transcription levels of gonad and brain aromatase genes, which encode the enzyme that converts androgens into estrogens, in the brains of dominant individuals. These results suggest that social rank reflects the blood Cortisol value, which in turn leads to sex differentiation by manipulating transcription of genes, including aromatase genes, in the brain.

Postoperative oral energy and protein intakes for an enhanced recovery after surgery program incorporating early enteral nutrition for pancreaticoduodenectomy: A retrospective study.

BACKGROUND: Although postoperative early oral feeding in the enhanced recovery after surgery (ERAS) program for pancreaticoduodenectomy (PD) is deemed safe, the assessment of oral intakes has been insufficient. This study aimed to investigate postoperative oral intakes and the effectiveness of an ERAS program incorporating early enteral nutrition (EN). METHODS: In total, 203 patients with PD were enrolled retrospectively. The first group (group E1; n = 61) comprised 11 ERAS care elements, whereas the second group (group E2; n = 106) comprised 19 elements. The control group (group C; n = 36) was managed using traditional care before ERAS was implemented. Postoperative energy and protein requirements were estimated at 25-30 kcal per kilogram of ideal body weight and 1.2-1.5 g per kilogram of ideal body weight, respectively, and were investigated along with the length of hospital stay (LOS). RESULTS: The oral energy and protein intakes from the diets in the ERAS groups at postoperative day 7 significantly increased compared with those in group C. Intakes in groups E1 and E2 were not significantly different and provided <30% of the requirements. However, the total intakes, which were compensated by EN, were maintained at >80% of the requirements. LOS was significantly shorter in groups E1 (31 days) and E2 (19 days) than in group C (52 days). CONCLUSIONS: Postoperative early oral energy and protein intakes of this modified ERAS program failed to meet the dietary requirements. However, early EN compensated for the shortages and contributed to the reduction of LOS.

Carbon-ion beam irradiation in combination with cisplatin effectively suppresses xenografted malignant pleural mesothelioma.

Malignant pleural mesothelioma (MPM) is a rare aggressive cancer. This study investigated the growth-inhibitory effects of the combination of carbon ion beam irradiation (IR) and cisplatin (CDDP) on MPM xenografts. Carbon-ion beam IR at 15 Gy effectively inhibited tumor growth and decreased the tumor volume more than 90% after 9 weeks. However, tumor regrowth was observed after 17 weeks. The combination of carbon-ion beam IR (15 Gy) and CDDP significantly suppressed tumor growth after 9 weeks, with tumor regression being observed for more than 18 weeks. In contrast, X-ray IR (30 Gy) alone or in combination with CDDP effectively suppressed tumor growth and decreased the tumor volume after 11 weeks, but tumor growth was observed after 15 weeks. Carbon-ion beam IR at 25 Gy resulted in complete tumor regression without tumor regrowth in the 20-week follow-up period. Histopathological analysis revealed that combination of carbon-ion beam IR and CDDP exerted effective cytotoxic effects on MPM xenograft tumor cells and significantly promoted tumor cell necrosis, cavitation, and fibrosis when compared with individual treatment with carbon-ion beam, X-ray IR, or CDDP. Immunohistochemical analysis revealed that the expression levels of tumor cell migration and invasion-related proteins such as CXCL12, MMP2 and MMP9 were not significantly affected upon low dose (15 Gy) carbon-ion beam IR alone or in combination with CDDP but were markedly upregulated upon treatment with CDDP alone relative to control. However, IR with a high dose (25 Gy) carbon-ion beam inhibited tumor growth without upregulating these proteins. In conclusion, the combination of IR with a low dose (15 Gy) carbon ion beam and CDDP effectively suppressed MPM tumor in vivo without significantly upregulating CXCL12, MMP2 and MMP9, suggesting that combination therapy of carbon ion beam IR and chemotherapy is a promising therapeutic strategy for MPM.

Three new species of Eimeria (Apicomplexa: Eimeriidae) from the Amami rabbit, Pentalagus furnessi (Mammalia: Leporidae).

The Amami rabbit, Pentalagus furnessi (Mammalia: Lagomorpha: Leporidae), is a relict and endangered species endemic to the Amami-Oshima and Tokunoshima Islands, located in southwestern Japan. Here, we described three new species of Eimeria (Apicomplexa: Eimeriidae) parasites detected from fecal samples of wild Amami rabbits. Eimeria furnessi n. sp., recorded in 21 (58.3%) samples, has ellipsoidal oocysts with two walls and micropyle, 26.0 × 16.6 µm, and elongate-ovoidal sporocysts, 13.1 × 6.3 µm, with Stieda body. Eimeria hilleri n. sp., recorded in 9 (25.0%) samples, has ellipsoidal oocysts with two walls and micropyle, 34.7 × 21.4 µm, and elongate-fusiform to elongate-ovoidal sporocysts, 15.7 × 8.3 µm, with Stieda and substieda bodies. Eimeria sagentae n. sp., recorded in 13 (36.1%) samples, has ellipsoidal oocysts with two walls and micropyle, 20.9 × 14.5 µm, and elongate-ovoidal sporocysts, 10.4 × 5.0 µm, with Stieda body. The three new species can be distinguished by the size and color of their oocysts. Further studies related to the pathogenicity of these parasites can improve the breeding and propagation procedures of the Amami rabbit.

Hippocampal Connectivity of the Presubiculum in the Common Marmoset (Callithrix jacchus).

The marmoset (a New World monkey) has recently received much attention as an experimental animal model; however, little is known about the connectivity of limbic regions, including cortical and hippocampal memory circuits, in the marmoset. Here, we investigated the neuronal connectivity of the marmoset, especially focusing on the connectivity between the hippocampal formation and the presubiculum, using retrograde and anterograde tracers (cholera toxin-B subunit and biotin dextran amine). We demonstrated the presence of a direct projection from the CA1 pyramidal cell layer to the deep layers of the presubiculum in the marmoset, which was previously identified in the rabbit brain, but not in the rat. We also found that the cells of origin of the subiculo-presubicular projections were localized in the middle part along the superficial-to-deep axis of the pyramidal cell layer of the distal subiculum in the marmoset, which was similar to that in both rats and rabbits. Our results suggest that, compared to the rat and rabbit brains, connections between the hippocampal formation and presubiculum are highly organized and characteristic in the marmoset brain.

A calcitonin receptor-expressing subregion of the medial preoptic area is involved in alloparental tolerance in common marmosets.

Like humans, common marmoset monkeys utilize family cooperation for infant care, but the neural mechanisms underlying primate parental behaviors remain largely unknown. We investigated infant care behaviors of captive marmosets in family settings and caregiver-infant dyadic situations. Marmoset caregivers exhibited individual variations in parenting styles, comprised of sensitivity and tolerance toward infants, consistently across infants, social contexts and multiple births. Seeking the neural basis of these parenting styles, we demonstrated that the calcitonin receptor-expressing neurons in the marmoset medial preoptic area (MPOA) were transcriptionally activated during infant care, as in laboratory mice. Further, site-specific neurotoxic lesions of this MPOA subregion, termed the cMPOA, significantly reduced alloparental tolerance and total infant carrying, while sparing general health and other social or nonsocial behaviors. These results suggest that the molecularly-defined neural site cMPOA is responsible for mammalian parenting, thus provide an invaluable model to study the neural basis of parenting styles in primates.