Identification and characterization of a novel α-haemolytic streptococci, Streptococcus parapneumoniae sp. nov., which caused bacteremia with pyelonephritis.

OBJECTIVES: We report a case of bacteremia with pyelonephritis in an adult male with an underlying disease caused by α-hemolytic streptococci. α-Hemolytic streptococci were isolated from blood, but it was challenging to identify its species. This study aimed to characterize the causative bacterium SP4011 and to elucidate its species. METHODS: The whole-genome sequence and biochemical characteristics of SP4011 were determined. Based on the genome sequence, phylogenetic analysis was performed with standard strains of each species of α-hemolytic streptococci. Digital DNA-DNA hybridization (dDDH) and average nucleotide identity (ANI) values were calculated. RESULTS: SP4011 showed optochin susceptibility and bile solubility, but did not react with pneumococcal omni antiserum. Phylogenetic analysis of the whole-genome sequence showed that SP4011 clustered with S. pneumoniae and S. pseodopneumoniae and was most closely related to S. pseodopneumoniae. Genomic analysis revealed that ANI and dDDH values between SP4011 and S. pseodopneumoniae were 94.0 % and 56.0 %, respectively, and between SP4011 and S. pneumoniae were 93.3 % and 52.2 %, respectively. Biochemical characteristics also showed differences between SP4011 and S. pseodopneumoniae and between SP4011 and S. pneumoniae. These results indicate that SP4011 is a novel species. CONCLUSION: Our findings indicate that SP4011 is a novel species of the genus Streptococcus. SP4011 has biochemical characteristics similar to S. pneumoniae, making it challenging to differentiate and requiring careful clinical diagnosis. This isolate was proposed to be a novel species, Streptococcus parapneumoniae sp. nov. The strain type is SP4011T (= JCM 36068T = KCTC 21228T).

Autopsy of tetanus with foot gangrene as portal of entry.

Non-traumatic chronic skin lesions are the second most common cause of tetanus. Herein, we describe an 85-year-old woman who presented with a chronically infected skin lesion. She developed tetanus while in hospital and died of respiratory failure, after refusing mechanical ventilation. Routine immunization against tetanus began in Japan during 1968; hence many people born before 1968 are unvaccinated. Mortality due to tetanus is high and the proportion with protective antibodies is low in older adults. Therefore, we recommend tetanus vaccination for older persons in Japan who have chronic skin lesions and have never been vaccinated.

Sexual fate of murine external genitalia development: Conserved transcriptional competency for male-biased genes in both sexes.

Testicular androgen is a master endocrine factor in the establishment of external genital sex differences. The degree of androgenic exposure during development is well known to determine the fate of external genitalia on a spectrum of female- to male-specific phenotypes. However, the mechanisms of androgenic regulation underlying sex differentiation are poorly defined. Here, we show that the genomic environment for the expression of male-biased genes is conserved to acquire androgen responsiveness in both sexes. Histone H3 at lysine 27 acetylation (H3K27ac) and H3K4 monomethylation (H3K4me1) are enriched at the enhancer of male-biased genes in an androgen-independent manner. Specificity protein 1 (Sp1), acting as a collaborative transcription factor of androgen receptor, regulates H3K27ac enrichment to establish conserved transcriptional competency for male-biased genes in both sexes. Genetic manipulation of MafB, a key regulator of male-specific differentiation, and Sp1 regulatory MafB enhancer elements disrupts male-type urethral differentiation. Altogether, these findings demonstrate conservation of androgen responsiveness in both sexes, providing insights into the regulatory mechanisms underlying sexual fate during external genitalia development.

Novel erectile analyses revealed augmentable penile Lyve-1, the lymphatic marker, expression.

Purpose: The pathophysiology of penis extends to erectile dysfunction (ED) to conditions including sexually transmitted diseases (STDs) and cancer. To date, there has been little research evaluating vascular drainage from the penis. We aimed to evaluate penile blood flow in vivo and analyze its possible relationship with the lymphatic maker. Materials and Methods: We established an in vivo system designed to assess the dynamic blood outflow from the corpus cavernosum (CC) by dye injection. To analyze lymphatic characteristics in the CC, the expression of Lyve-1, the key lymphatic endothelium marker, was examined by the in vitro system and lipopolysaccharide (LPS) injection to mimic the inflammatory conditions. Results: A novel cavernography methods enable high-resolution morphological and functional blood drainage analysis. The expression of Lyve-1 was detected along the sinusoids. Furthermore, its prominent expression was also observed after penile LPS injection and in the erectile condition. Conclusions: The current in vivo system will potentially contribute to the assessment of penile pathology from a novel viewpoint. In addition, current analyses revealed inducible Lyve-1 expression for LPS injection and the erection state, which requires further analyses on penile lymphatic system.

Increased infiltration of CD4+ T cell in the complement deficient lymphedema model.

BACKGROUND: Lymphedema is an intractable disease that can be caused by injury to lymphatic vessels, such as by surgical treatments for cancer. It can lead to impaired joint mobility in the extremities and reduced quality of life. Chronic inflammation due to infiltration of various immune cells in an area of lymphedema is thought to lead to local fibrosis, but the molecular pathogenesis of lymphedema remains unclear. Development of effective therapies requires elucidation of the immunological mechanisms involved in the progression of lymphedema. The complement system is part of the innate immune system which has a central role in the elimination of invading microbes and acts as a scavenger of altered host cells, such as apoptotic and necrotic cells and cellular debris. Complement-targeted therapies have recently been clinically applied to various diseases caused by complement overactivation. In this context, we aimed to determine whether complement activation is involved in the development of lymphedema. RESULTS: Our mouse tail lymphedema models showed increased expression of C3, and that the classical or lectin pathway was locally activated. Complement activation was suggested to be involved in the progression of lymphedema. In comparison of the C3 knockout (KO) mouse lymphedema model and wild-type mice, there was no difference in the degree of edema at three weeks postoperatively, but the C3 KO mice had a significant increase of TUNEL+ necrotic cells and CD4+ T cells. Infiltration of macrophages and granulocytes was not significantly elevated in C3 KO or C5 KO mice compared with in wild-type mice. Impaired opsonization and decreased migration of macrophages and granulocytes due to C3 deficiency should therefore induce the accumulation of dead cells and may lead to increased infiltration of CD4+ T cells. CONCLUSIONS: Vigilance for exacerbation of lymphedema is necessary when surgical treatments have the potential to injure lymphatic vessels in patients undergoing complement-targeted therapies or with complement deficiency. Future studies should aim to elucidate the molecular mechanism of CD4+ T cell infiltration by accumulated dead cells.

The role of p63 in embryonic external genitalia outgrowth in mice.

Embryonic external genitalia (genital tubercle [GT]) protrude from the cloaca and outgrow as cloacal development progresses. Individual gene functions and knockout phenotypes in GT development have been extensively analyzed; however, the interactions between these genes are not fully understood. In this study, we investigated the role of p63, focusing on its interaction with the Shh-Wnt/Ctnnb1-Fgf8 pathway, a signaling network that is known to play a role in GT outgrowth. p63 was expressed in the epithelial tissues of the GT at E11.5, and the distal tip of the GT predominantly expressed the ΔNp63α isoform. The GTs in p63 knockout embryos had normal Shh expression, but CTNNB1 protein and Fgf8 gene expression in the distal urethral epithelium was decreased or lost. Constitutive expression of CTNNB1 in p63-null embryos restored Fgf8 expression, accompanied by small bud structure development; however, such bud structures could not be maintained by E13.5, at which point mutant GTs exhibited severe abnormalities showing a split shape with a hemorrhagic cloaca. Therefore, p63 is a key component of the signaling pathway that triggers Fgf8 expression in the distal urethral epithelium and contributes to GT outgrowth by ensuring the structural integrity of the cloacal epithelia. Altogether, we propose that p63 plays an essential role in the signaling network for the development of external genitalia.

Effectiveness of oral cephalexin in antibiotic-course completion for methicillin-susceptible Staphylococcus aureus-induced bacteremic vertebral osteomyelitis.

BACKGROUND: Methicillin-susceptible Staphylococcus aureus (MSSA) is the most common causative microorganism of pyogenic vertebral osteomyelitis (PVO). Although oral antimicrobial therapy with first-generation cephalosporins can treat MSSA infection, data on PVO are scarce. This study evaluated the treatment efficacy of cephalexin as oral antibiotic therapy for MSSA-induced PVO. METHODS: This retrospective study included adult patients treated with oral cephalexin as the completing treatment for PVO with MSSA bacteremia from 2012 to 2020. Treatment effectiveness of cephalexin was evaluated by comparing improvement (5-point scale; score ≥ 4/5 indicates treatment success) in symptoms and laboratory and imaging results between intravenous antimicrobial and oral cephalexin treatment. RESULTS: Among 15 participants (8 [53%] women; median [interquartile range, IQR], age 75 [67.5-80.5] years; Charlson Comorbidity Index 2 [0-4]), 10 (67%) had lumbar spine lesions, 12 (80%) had spinal abscesses, and 4 (27%) had remote abscesses; no patients had concomitant endocarditis. In 11 patients with normal renal function, cephalexin 1,500-2,000 mg/day was administered. Five patients (33%) underwent surgery. Median (IQR; range) duration (days) of intravenous antibiotics, cephalexin, and total treatment was 36 (32-61; 21-86), 29 (19-82; 8-251), and 86 (59-125; 37-337), respectively. Cephalexin had an 87% treatment success rate without recurrence during a median follow-up of 119 (IQR, 48.5-350) days. CONCLUSIONS: In patients with MSSA bacteremia and PVO, antibiotic treatment completion with cephalexin is a reasonable option, even in cases with spinal abscess, if at least 3 weeks of effective intravenous antimicrobial therapy is provided.

A case of prosthetic joint infection caused by Mycobacterium tuberculosis complicated secondary bacterial infection after knee joint replacement surgery.

Mycobacterium tuberculosis (M. tuberculosis) is a rare cause of prosthetic joint infection (PJI). Previous studies have reported that many cases of PJI caused by M. tuberculosis have no medical history of active tuberculosis (TB) or other localization, which contributes to diagnostic difficulties. Furthermore, owing to the limited number of studies on treatment, appropriate treatment strategies, such as the duration of anti-tuberculosis (anti-TB) drugs and surgical indications, remain unclear. We report a case of PJI caused by M. tuberculosis and secondary pyogenic arthritis caused by Staphylococcus aureus and Streptococcus dysgalactiae in a 67-year-old man after knee joint replacement surgery in Japan, which was a moderately endemic country until 2020 and a low endemic country since 2021. Although he had no past medical history or close contact with TB, he was diagnosed with PJI caused by M. tuberculosis, following the culture of a synovectomy specimen. He underwent two-stage surgery and was treated with anti-TB drugs for a total of 12 months and recovered without recurrence. Based on our case and previous studies, there are three points of clinical significance for PJI caused by M. tuberculosis. First, about one year of anti-TB drugs with two staged joint revision resulted in a good course of treatment. Second, surgical treatment might be considered in cases complicated by secondary bacterial infection. Third, because the diagnosis of PJI caused by M. tuberculosis is difficult, TB should be considered in the differential diagnosis of routine bacterial culture-negative PJI, especially in endemic areas.

Clinical outcomes and epidemiological characteristics of bacteremia in the older Japanese population.

BACKGROUND: The characteristics and clinical consequences of bacteremia in older people, who are highly susceptible to infections, need to be clarified. This study aimed to determine the epidemiological characteristics, prognosis, and predictors of 7-day mortality in patients with community-acquired (CA), healthcare-associated (HCA), and hospital-onset (HO) bacteremia in older adults aged ≥65 years. METHODS: Patients aged ≥65 years with positive blood cultures between April 1, 2015, and March 31, 2018, were divided into three groups: pre-old (65-74 years), old (75-89 years), and super-old (≥90 years). Characteristics based on medical exposure, including CA, HCA, and HO, were also compared and factors related to mortality were identified. RESULTS: Overall, 1716 episodes of bacteremia were identified in 1415 patients. Of the 1211 episodes without contamination, 32.8%, 54.3%, and 12.9% occurred in pre-old, old, and super-old patients. Central line-associated bloodstream infections were more common in pre-old patients and urinary tract infections in the old and super-old. The 7-day mortality rates in the pre-old, old, and super-old groups were 7.4%, 5.8%, and 14.2% (P = 0.002), respectively. Multivariable logistic regression showed that super-old age (adjusted odds ratio, aOR: 2.09 [1.13-3.88], P = 0.019) and HO bacteremia (aOR: 1.97 [1.18-3.28], P = 0.010) were independent risk factors for 7-day mortality. Infectious disease consultation had a protective effect on 7-day mortality (aOR: 0.59 [0.35-0.99], P = 0.047). CONCLUSIONS: The epidemiology of bacteremia differs among older people; thus, they should not be treated as a single entity. A careful approach is needed for the optimal management of bacteremia in these vulnerable patients.

Potential usage of anterior nasal sampling in clinical practice with three rapid antigen tests for SARS-CoV-2.

INTRODUCTION: Anterior nasal sampling (AN) might be more convenient for patients than NP sampling to diagnose coronavirus disease. This study investigated the feasibility of rapid antigen tests for AN sampling, and the factors affecting the test accuracy. METHODS: This single-center prospective study evaluated one qualitative (ESP) and two quantitative (LUMI and LUMI-P) rapid antigen tests using AN and NP swabs. Symptomatic patients aged 20 years or older, who were considered eligible for reverse-transcription quantitative polymerase chain reaction using NP samples within 9 days of onset were recruited. Sensitivity, specificity, and positive and negative concordance rates between AN and NP samples were assessed for the rapid antigen tests. We investigated the characteristics that affected the concordance between AN and NP sampling results. RESULTS: A total of 128 cases were recruited, including 28 positive samples and 96 negative samples. The sensitivity and specificity of AN samples using ESP were 0.81 and 1.00, while those of NP samples were 0.94 and 1.00. The sensitivity of AN and NP samples was 0.91 and 0.97, respectively, and specificity was 1.00, for both LUMI and LUMI-P. The positive concordance rates of AN to NP sampling were 0.87, 0.94, and 0.85 for ESP, LUMI, and LUMI-P, respectively. No factor had a significant effect on the concordance between the sampling methods. CONCLUSIONS: ESP, LUMI, and LUMI-P showed practical diagnostic accuracy for AN sampling compared to NP sampling. There was no significant factor affecting the concordance between AN and NP sampling for these rapid antigen tests.

Emerging structural and pathological analyses on the erectile organ, corpus cavernous containing sinusoids.

Background: The corpus cavernosum (CC) containing sinusoids plays fundamental roles for erection. Analysis of pathological changes in the erectile system is studied by recent experimental systems. Various in vitro models utilizing genital mesenchymal-derived cells and explant culture systems are summarized. Methods: 3D reconstruction of section images of murine CC was created. Ectopic chondrogenesis in aged mouse CC was shown by a gene expression study revealing the prominent expression of Sox9. Various experimental strategies utilizing mesenchyme-derived primary cells and tissue explants are introduced. Main Findings: Possible roles of Sox9 in chondrogenesis and its regulation by several signals are suggested. The unique character of genital mesenchyme is shown by various analyses of external genitalia (ExG) derived cells and explant cultures. Such strategies are also applied to the analysis of erectile contraction/relaxation responses to many signals and aging process. Conclusion: Erectile dysfunction (ED) is one of the essential topics for the modern aged society. More comprehensive studies are necessary to reveal the nature of the erectile system by combining multiple cell culture strategies.

Two Cases of Breakthrough Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Infections Caused by the Omicron Variant (B.1.1.529 Lineage) in International Travelers to Japan.

In November 2021, the World Health Organization designated a new severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variant of concern, Omicron (PANGO lineage B.1.1.529). We report on the first 2 cases of breakthrough coronavirus disease 2019 (COVID-19) caused by Omicron in Japan among international travelers returning from the country with undetected infection. The spread of infection by Omicron were considered.

Multisystem Inflammatory Syndrome in Adult after First Dose of mRNA Vaccine.

A 32-year-old man in Japan experienced respiratory failure after receiving the first dose of coronavirus disease (COVID-19) vaccine. He was treated with noninvasive ventilation and corticosteroids. Serologic test results suggested previous COVID-19; therefore, he received a diagnosis of multisystem inflammatory syndrome. COVID-19 vaccination could be a trigger for this condition.

Systemic corticosteroid as an adjunctive treatment for lower respiratory tract infection in children with severe motor and intellectual disabilities.

INTRODUCTION: Children with severe motor and intellectual disabilities (SMID) are susceptible to severe lower respiratory tract infection (LTRI). As SMID patients are prone to develop recurrent wheezing and are often diagnosed with bronchial asthma, they frequently receive systemic corticosteroids as an adjunctive treatment for LRTIs. However, the efficacy of corticosteroid therapy for LTRIs in SMID children is unclear. We investigated whether or not corticosteroid therapy was associated with better clinical outcomes for SMID children with LRTIs. METHODS: Our retrospective study enrolled 217 SMID children 1-15 years old hospitalized for LTRIs. We compared the clinical characteristics and outcomes between patients with and without corticosteroid therapy. RESULTS: Of the 217 patients, 29 (13.3%) received corticosteroid therapy. The proportion of patients with a history of bronchial asthma was higher and LRTI was more severe in patients with corticosteroid therapy than in those without the therapy. The length of hospital stay (LOHS) was significantly longer in patients with corticosteroid therapy (median 13 days) than in those without corticosteroid therapy (median 9 days) (P = 0.02). The same tendency was shown for the LOHS in patients with severe or moderate LRTI, although not to a significant extent. CONCLUSION: Systemic corticosteroid therapy was not associated with better clinical outcomes in SMID children with LRTIs, even if the patients suffer from severe LRTIs. Corticosteroids should be used cautiously for LRTIs in SMID children because bronchial asthma is likely to be overdiagnosed in these children.

Predictors of silent hypoxia in hospitalized patients with COVID-19 in Japan.

INTRODUCTION: Silent hypoxia (SH) is common in patients with coronavirus disease (COVID-19) in Japan and other countries. Early identification of SH is important as more treatment options for COVID-19 have become available. This study aimed to identify predictors of SH using a nationwide COVID-19 registry of hospitalized patients. METHODS: Adult patients who were admitted to hospital with COVID-19 between January 2020 and June 2021 and who were hypoxic on admission (SpO2: 70-93%), not transferred from another facility, and who did not have disturbance of consciousness, confusion, or dementia, were included. SH was defined as hypoxia in the absence of shortness of breath/dyspnea upon admission. Predictors of SH were identified using univariable and multivariable logistic regression. RESULTS: The study included 1904 patients, of whom 990 (52%) satisfied the criteria for SH. Compared to patients without SH, patients with SH were older, more likely to be female, and had a slightly higher SpO2 on admission. Compared to patients without SH, patients with SH had a lower prevalence of chronic lung disease (CLD) other than chronic obstructive pulmonary disease (COPD), asthma, and obesity. Multivariable analysis revealed that the independent predictors of SH were older age, a shorter interval from symptom onset to admission, higher SpO2, and an absence of CLD or COPD. CONCLUSIONS: The absence of underlying lung disease and older age were important predictors of SH. The results of this study, which is the largest such study reported to date in Japan, may help clarify the mechanism of SH.

Factors associated with high antibody titer following coronavirus disease among 581 convalescent plasma donors: A single-center cross-sectional study in Japan.

INTRODUCTION: The ability to predict which patients with a history of coronavirus disease (COVID-19) will exhibit a high antibody titer is necessary for more efficient screening of potential convalescent plasma donors. We aimed to identify factors associated with a high immunoglobulin G (IgG) titer in Japanese convalescent plasma donors after COVID-19. METHODS: This cross-sectional study included volunteers undergoing screening for convalescent plasma donation after COVID-19. Serum anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) S-protein IgG antibodies were measured using a high-sensitivity chemiluminescence enzyme immunoassay. RESULTS: IgG antibodies were measured in 581 patients, 534 of whom had full information of selected independent variables. Multiple linear regression analysis revealed that increasing age (1.037 [1,025, 1.048]), days from symptom onset to sampling (0.997 [0.995, 0.998]), fever (1.664 [1.226, 2.259]), systemic corticosteroid use during SARS-CoV-2 infection (2.382 [1.576, 3.601]), and blood type AB (1.478 [1.032, 2.117]) predict antibody titer. CONCLUSION: Older participants, those who experienced fever during infection, those treated with systemic corticosteroids during infection, those from whom samples were obtained earlier after symptom onset, and those with blood type AB are the best candidates for convalescent plasma donation. Therefore, these factors should be incorporated into the screening criteria for convalescent plasma donation after SARS-CoV-2 infection.

Fatty liver on computed tomography scan on admission is a risk factor for severe coronavirus disease.

OBJECTIVES: To alleviate the overflow of coronavirus disease 2019 (COVID-19) patients in hospitals, less invasive and simple criteria are required to triage the patients. We evaluated the relationship between COVID-19 severity and fatty liver on plain computed tomography (CT) scan performed on admission. METHODS: In this retrospective cohort study, we considered all COVID-19 patients at a large tertiary care hospital between January 31 and August 31, 2020. COVID-19 severity was categorized into severe (moderate and severe) and non-severe (asymptomatic and mild) groups, based on the Japanese National COVID-19 guidelines. Fatty liver was detected on plain CT scan. Multivariate logistic regression analysis was performed to evaluate factors associated with severe COVID-19. RESULTS: Of 222 patients (median age: 52 years), 3.2%, 58.1%, 20.7%, and 18.0% presented with asymptomatic, mild, moderate, and severe COVID-19, respectively. Although 59.9% had no fatty liver on plain CT, mild, moderate, and severe fatty liver occurred in 13.1%, 18.9%, and 8.1%, respectively. Age and presence of fatty liver were significantly associated with severe COVID-19. CONCLUSION: Our study showed that fatty liver on plain CT scan on admission can become a risk factor for severe COVID-19. This finding may help clinicians to easily triage COVID-19 patients.

Establishment of mouse line showing inducible priapism-like phenotypes.

Purpose: Penile research is expected to reveal new targets for treatment and prevention of the complex mechanisms of its disorder including erectile dysfunction (ED). Thus, analyses of the molecular processes of penile ED and continuous erection as priapism are essential issues of reproductive medicine. Methods: By performing mouse N-ethyl-N-nitrosourea mutagenesis and exome sequencing, we established a novel mouse line displaying protruded genitalia phenotype (PGP; priapism-like phenotype) and identified a novel Pitpna gene mutation for PGP. Extensive histological analyses on the Pitpna mutant and intracavernous pressure measurement (ICP) and liquid chromatography-electrospray ionization tandem mass spectrometry (LC-ESI/MS)/MS analyses were performed. Results: We evaluated the role of phospholipids during erection for the first time and showed the mutants of inducible phenotypes of priapism. Moreover, quantitative analysis using LC-ESI/MS/MS revealed that the level of phosphatidylinositol (PI) was significantly lower in the mutant penile samples. These results imply that PI may contribute to penile erection by PITPα. Conclusions: Our findings suggest that the current mutant is a mouse model for priapism and abnormalities in PI signaling pathways through PITPα may lead to priapism providing an attractive novel therapeutic target in its treatment.

Impact of Body Weight Gain on the Incidence of Nonalcoholic Fatty Liver Disease in Nonobese Japanese Individuals.

INTRODUCTION: We aimed to investigate the effect of recent short-term weight gain on the incidence of nonalcoholic fatty liver disease (NAFLD) in nonobese (body mass index < 25 kg/m) participants. METHODS: This retrospective cohort study included nonobese individuals who participated in an annual health checkup between 2008 and 2018 in Tokyo, Japan. We estimated the multivariable adjusted hazard ratio for the development of NAFLD diagnosed via ultrasound after a 3-kg unit gain in weight measured at a 2-year landmark time point postbaseline. Multivariable adjustments included weight change from the age of 20 and other relevant confounding factors. Sensitivity analyses using additional landmark time points at 1, 3, 4, and 5 years postbaseline and time-dependent Cox proportional hazards regressions were performed. RESULTS: Among the 27,064 nonobese participants (142,699 person years of follow-up), 2,895 were diagnosed with NAFLD. Approximately 90% of the patients with NAFLD maintained their nonobese status before disease diagnosis. The adjusted hazard ratio for the development of NAFLD (for a 3-kg unit of weight gain) at the 2-year landmark time point postbaseline was 1.60 (95% confidence interval, 1.46-1.76) in nonobese men and 1.66 (95% confidence interval, 1.51-1.83) in nonobese women. This association was maintained in the sensitivity analyses. DISCUSSION: Recent short-term weight gain is an independent risk factor for NAFLD development in nonobese men and women. Clinicians should be mindful of the association between weight gain and NAFLD onset, even in the nonobese population.

Development of Surgical and Visualization Procedures to Analyze Vasculatures by Mouse Tail Edema Model.

BACKGROUND: Because of the high frequency of chronic edema formation in the current "aged" society, analyses and detailed observation of post-surgical edema are getting more required. Post-surgical examination of the dynamic vasculature including L.V. (Lymphatic Vasculature) to monitor edema formation has not been efficiently performed. Hence, procedures for investigating such vasculature are essential. By inserting transparent sheet into the cutaneous layer of mouse tails as a novel surgery model (the Tail Edema by Silicone sheet mediated Transparency protocol; TEST), the novel procedures are introduced and analyzed by series of histological analyses including video-based L.V. observation and 3D histological reconstruction of vasculatures in mouse tails. RESULTS: The dynamic generation of post-surgical main and fine (neo) L.V. connective structure during the edematous recovery process was visualized by series of studies with a novel surgery model. Snapshot images taken from live binocular image recording for TEST samples suggested the presence of main and elongating fine (neo) L.V. structure. After the ligation of L.V., the enlargement of main L.V. was confirmed. In the case of light sheet fluorescence microscopy (LSFM) observation, such L.V. connections were also suggested by using transparent 3D samples. Finally, the generation of neo blood vessels particularly in the region adjacent to the silicone sheet and the operated boundary region was suggested in 3D reconstruction images. However, direct detection of elongating fine (neo) L.V. was not suitable for analysis by such LSFM and 3D reconstruction procedures. Thus, such methods utilizing fixed tissues are appropriate for general observation for the operated region including of L.V. CONCLUSIONS: The current surgical procedures and analysis on the post-surgical status are the first case to observe vasculatures in vivo with a transparent sheet. Systematic analyses including the FITC-dextran mediated snap shot images observation suggest the elongation of fine (neo) lymphatic vasculature. Post-surgical analyses including LSFM and 3D histological structural reconstruction, are suitable to reveal the fixed structures of blood and lymphatic vessels formation.