Computational Fluid Dynamics Using a Porous Media Setting Predicts Outcome after Flow-Diverter Treatment.

BACKGROUND AND PURPOSE: Knowledge about predictors of the outcome of flow-diverter treatment is limited. The aim of this study was to predict the angiographic occlusion status after flow-diverter treatment with computational fluid dynamics using porous media modeling for decision-making in the treatment of large wide-neck aneurysms. MATERIALS AND METHODS: A total of 27 patients treated with flow-diverter stents were retrospectively analyzed through computational fluid dynamics using pretreatment patient-specific 3D rotational angiography. These patients were classified into no-filling and contrast-filling groups based on the O'Kelly-Marotta scale. The patient characteristics, morphologic variables, and hemodynamic parameters were evaluated for understanding the outcomes of the flow-diverter treatment. RESULTS: The patient characteristics and morphologic variables were similar between the 2 groups. Flow velocity, wall shear stress, shear rate, modified aneurysmal inflow rate coefficient, and residual flow volume were significantly lower in the no-filling group. A novel parameter, called the normalized residual flow volume, was developed and defined as the residual flow volume normalized by the dome volume. The receiver operating characteristic curve analyses demonstrated that the normalized residual flow volume with an average flow velocity of ≥8.0 cm/s in the aneurysmal dome was the most effective in predicting the flow-diverter treatment outcomes. CONCLUSIONS: It was established in this study that the hemodynamic parameters could predict the angiographic occlusion status after flow-diverter treatment.

Intracranial Subdural Hematoma after Lumbar Spine Surgery: A Case Report.

Intracranial subdural hematoma following lumbar surgery is a devastating but rare complication. It has been implicated due to intracranial hypotension secondary to persistent cerebrospinal fluid leakage. The resultant drop in intracranial pressure presumably causes traction and tearing of venous structures. Patients typically present with postural headaches. However, other symptoms of subdural hematoma, intracranial hypotension and cerebrospinal fluid leak must also be cautioned.

Should sagittal osteotomy line be parallel to tibial posterior slope in high tibial osteotomy?

BACKGROUND: The reason why the osteotomy line in the sagittal view should be parallel to the medial tibial posterior slope in open wedge high tibial osteotomy (OWHTO) remains unclear. In addition, previous study reported that a posterolateral hinge position led to an increase in tibial posterior slope (TPS) after OWHTO. Our aims were to examine the relationships between angles among the tibial plateau and osteotomy planes or the hinge point and the change in TPS, and the location of the hinge position after OWHTO using three-dimensional computed tomography (3DCT). We hypothesized that the sagittal angle between the tibial plateau and osteotomy planes with an anterior-widening proximal tibial fragment resulted in increased TPS, and the hinge position located posterolaterally. METHODS: Preoperative planning anticipated a weight-bearing line ratio of 62% on the radiograph. The anterior gap was 67% of the posterior gap in OWHTO. We identified the tibial plateau and upper and lower osteotomy planes on 3DCT of 82 patients with symptomatic medial osteoarthritic knee after OWHTO. The osteotomy plane angles between the tibial plateau and upper osteotomy planes, and opening gap angles between both osteotomy planes in the coronal and sagittal views were measured. The anteroposterior (AP) and lateral hinge position was displayed as a percentage on the upper osteotomy plane. We assessed the relationships among them. RESULTS: The TPS significantly increased after OWHTO (p = 0.002). There was no significant difference between the sagittal osteotomy plane angle and the change in TPS. The sagittal opening gap angle and the AP hinge position ratio were significantly correlated with the change in the TPS (r = 0.477 p < 0.001 and r = - 0.342, p = 0.002, respectively). The hinge position was located a mean of 16.0% from the lateral and 48.6% from the posterior tibial edge in the upper osteotomy plane. CONCLUSIONS: Contrary to our expectation, the osteotomy plane did not need to be parallel to the tibial plateau plane in the sagittal view. However, the osteotomy gap should be rectangular in the sagittal view. The hinge position located nearly in the center of the sagittal view.

Chloride ion transport and overexpression of TMEM16A in a guinea-pig asthma model.

BACKGROUND: TMEM16A, a Ca-activated Cl channel, regulates various physiological functions such as mucin secretion. However, the role of TMEM16A in hyper-secretion in asthma is not fully understood. OBJECTIVE: The aim of this study is to evaluate Cl ion transport via TMEM16A and determine the localization of TMEM16A in a guinea-pig asthma model. METHODS: Guinea-pigs were sensitized with ovalbumin (OVA) i.p. on Days 1 and 8. On Day 22, we assessed OVA challenge-induced Cl ion transport in the sensitized tracheas ex vivo in an Ussing chamber, compared with the non-sensitized tracheas. We then examined the effect of T16Ainh-A01, a TMEM16A inhibitor, on the increase in Cl ion transport. The tracheal epithelium was immunostained with an anti-TMEM16A antibody. Epithelial cells from guinea-pig tracheas were cultured at the air-liquid interface in the presence of IL-13 for in vitro study. We studied the effect of TMEM16A inhibitors on Ca-dependent agonist, uridine triphosphate (UTP)-induced increases in Cl ion transport in the cultured cells. The cells were immunostained with an anti-TMEM16A antibody, an anti-MUC5AC antibody and an anti-α-tubulin antibody. RESULTS: OVA challenge induced an increase in short circuit current within 1 min in the OVA-sensitized tracheas but not in the non-sensitized tracheas, which was inhibited by pretreatment of T16Ainh-A01. Sensitized tracheas showed goblet cell metaplasia with more positive TMEM16A immunostaining, particularly in the apical portion compared with the non-sensitized tracheas. The in vitro UTP-induced increase in Cl ion transport was strongly inhibited by pretreatment with T16Ainh-A01, benzbromarone, and niflumic acid. TMEM16A was positively immunostained at the apical portion and in the MUC5AC-positive area in IL-13-induced goblet cell metaplasia. CONCLUSIONS: Antigen challenge and Ca-dependent agonist treatment increased Cl ion transport via the overexpression of TMEM16A in goblet cell metaplasia in a guinea-pig asthma model. TMEM16A inhibitors may be useful for the treatment of hyper-secretion in asthma.

Automatic generation of smart earthquake-resistant building system: Hybrid system of base-isolation and building-connection.

A base-isolated building may sometimes exhibit an undesirable large response to a long-duration, long-period earthquake ground motion and a connected building system without base-isolation may show a large response to a near-fault (rather high-frequency) earthquake ground motion. To overcome both deficiencies, a new hybrid control system of base-isolation and building-connection is proposed and investigated. In this new hybrid building system, a base-isolated building is connected to a stiffer free wall with oil dampers. It has been demonstrated in a preliminary research that the proposed hybrid system is effective both for near-fault (rather high-frequency) and long-duration, long-period earthquake ground motions and has sufficient redundancy and robustness for a broad range of earthquake ground motions.An automatic generation algorithm of this kind of smart structures of base-isolation and building-connection hybrid systems is presented in this paper. It is shown that, while the proposed algorithm does not work well in a building without the connecting-damper system, it works well in the proposed smart hybrid system with the connecting damper system.

Long-Term Persistence of Donor Alveolar Macrophages in Human Lung Transplant Recipients That Influences Donor-Specific Immune Responses.

Steady-state alveolar macrophages (AMs) are long-lived lung-resident macrophages with sentinel function. Evidence suggests that AM precursors originate during embryogenesis and populate lungs without replenishment by circulating leukocytes. However, their presence and persistence are unclear following human lung transplantation (LTx). Our goal was to examine donor AM longevity and evaluate whether AMs of recipient origin seed the transplanted lungs. Origin of AMs was accessed using donor-recipient HLA mismatches. We demonstrate that 94-100% of AMs present in bronchoalveolar lavage (BAL) were donor derived and, importantly, AMs of recipient origin were not detected. Further, analysis of BAL cells up to 3.5 years post-LTx revealed that the majority of AMs (>87%) was donor derived. Elicitation of de novo donor-specific antibody (DSA) is a major post-LTx complication and a risk factor for development of chronic rejection. The donor AMs responded to anti-HLA framework antibody (Ab) with secretion of inflammatory cytokines. Further, in an experimental murine model, we demonstrate that adoptive transfer of allogeneic AMs stimulated humoral and cellular immune responses to alloantigen and lung-associated self-antigens and led to bronchiolar obstruction. Therefore, donor-derived AMs play an essential role in the DSA-induced inflammatory cascade leading to obliterative airway disease of the transplanted lungs.

A genome-wide association study of periodontitis in a Japanese population.

Periodontitis is a multifactorial disease in which bacterial, lifestyle, and genetic factors are involved. Although previous genetic association studies identified several susceptibility genes for periodontitis in European populations, there is little information for Asian populations. Here, we conducted a genome-wide association study and a replication study consisting of 2,760 Japanese periodontitis patients and 15,158 Japanese controls. Although single-nucleotide polymorphisms that surpassed a stringent genome-wide significance threshold (P < 5 × 10(-8)) were not identified, we found 2 suggestive loci for periodontitis: KCNQ5 on chromosome 6q13 (rs9446777, P = 4.83 × 10(-6), odds ratio = 0.82) and GPR141-NME8 at chromosome 7p14.1 (rs2392510, P = 4.17 × 10(-6), odds ratio = 0.87). A stratified analysis indicated that the GPR141-NME8 locus had a strong genetic effect on the susceptibility to generalized periodontitis in Japanese individuals with a history of smoking. In conclusion, this study identified 2 suggestive loci for periodontitis in a Japanese population. This study should contribute to a further understanding of genetic factors for enhanced susceptibility to periodontitis.

Relative incidences and outcomes of Clostridium difficile infection following transplantation of unrelated cord blood, unrelated bone marrow, and related peripheral blood in adult patients: a single institute study.

BACKGROUND: Clostridium difficile is a major cause of nosocomial diarrhea. The incidence and prognosis of C. difficile-associated diarrhea (CDAD) has not yet been assessed in adult patients after unrelated cord blood transplantation (uCBT). METHODS: The medical records of 135 adult unrelated cord blood transplant recipients were reviewed retrospectively to investigate the clinical features of CDAD after uCBT. These data were compared to medical records of 39 unrelated bone marrow transplant recipients and 27 related peripheral blood stem cell transplant recipients as controls. RESULTS: A total of 17 recipients developed CDAD, with onset occurring at a median of 22 days (range, 0-56 days) after transplantation. Among the unrelated cord blood transplant recipients, 11 (9%) developed CDAD. These results were comparable with those of CDAD after unrelated bone marrow transplantation (uBMT) (2/39, 6%) and related peripheral blood stem cell transplantation (rPBSCT) (4/27, 16%) (P=0.37). Fifteen of the infected recipients were successfully treated with oral metronidazole, vancomycin, or cessation of antibiotics. The remaining 2 recipients who developed CDAD after uCBT died of other causes. The development of CDAD did not negatively affect overall survival after uCBT. CONCLUSIONS: These data indicate that the incidence and prognosis of CDAD after uCBT are comparable with those after uBMT and rPBSCT.

Multiple cytokine-producing testicular malignant lymphoma with clinical symptoms resembling infectious signs.

We present the case of a 64-year-old male with painful swelling of the bilateral testes and epididymides, high fever, leukocytosis, and an elevated C-reactive protein (CRP) level. This is the first case report of testicular diffuse large B-cell lymphoma, not otherwise specified (DLBCL, NOS) immunostained for multiple cytokines and their receptors, which clearly demonstrates that tumor cells express multiple cytokines [interleukin-6 (IL-6) and granulocyte colony-stimulating factor (G-CSF)] and their receptors [IL-6 receptor (IL-6R) and G-CSF receptor (G-CSFR)]. The clinical course showed that the reduction in tumor size was accompanied by a corresponding improvement in clinical symptoms and peripheral blood findings. Such clinical investigation may lead clinicians to misdiagnose inflammatory disease rather than neoplastic disease. Recognizing this paraneoplastic phenomenon associated with some cases of testicular DLBCL, NOS is important. In addition, this case suggests that the growth of tumor cells may be promoted through autocrine mechanisms of IL-6 and G-CSF, which are produced by tumor cells. The possibility that these cytokines can be produced by tumor cells and can accelerate tumor proliferation should be considered to be a cause of severe clinical symptoms, an aggressive clinical course, and an indication of the necessity of treatment. Certain cytokines may be used as tumor markers in some cases of DLBCL, NOS.

Effects of intravenous administration of umbilical cord blood CD34(+) cells in a mouse model of neonatal stroke.

Neonatal stroke occurs in approximately 1/4000 live births and results in life-long neurological impairments: e.g., cerebral palsy. Currently, there is no evidence-based specific treatment for neonates with stroke. Several studies have reported the benefits of umbilical cord blood (UCB) cell treatment in rodent models of neonatal brain injury. However, all of the studies examined the effects of administering either the UCB mononuclear cell fraction or UCB-derived mesenchymal stem cells in neonatal rat models. The objective of this study was to examine the effects of human UCB CD34(+) cells (hematopoietic stem cell/endothelial progenitor cells) in a mouse model of neonatal stroke, which we recently developed. On postnatal day 12, immunocompromized (SCID) mice underwent permanent occlusion of the left middle cerebral artery (MCAO). Forty-eight hours after MCAO, human UCB CD34(+) cells (1×10(5)cells) were injected intravenously into the mice. The area in which cerebral blood flow (CBF) was maintained was temporarily larger in the cell-treated group than in the phosphate-buffered saline (PBS)-treated group at 24h after treatment. With cell treatment, the percent loss of ipsilateral hemispheric volume was significantly ameliorated (21.5±1.9%) compared with the PBS group (25.6±5.1%) when assessed at 7weeks after MCAO. The cell-treated group did not exhibit significant differences from the PBS group in either rotarod (238±46s in the sham-surgery group, 175±49s in the PBS group, 203±54s in the cell-treated group) or open-field tests. The intravenous administration of human UCB CD34(+) cells modestly reduced histological ischemic brain damage after neonatal stroke in mice, with a transient augmentation of CBF in the peri-infarct area.

An approach for plasma cell myeloma diagnosis by two-color flow cytometry based on kappa/lambda ratios of CD38-gated plasma cells.

Criteria from the World Health Organization (WHO) are commonly used to diagnose plasma cell myeloma (PCM), but they are complex and require several laboratory parameters. To differentiate reactive plasmacytosis from clonal plasma cell neoplasms, such as PCM, it is important to accurately determine the expression of the cytoplasmic immunoglobulin (cIg) light chain (LC). Through retrospective analyses, we selected the patients with PCM, and analyzed records of 52 PCM patients, who underwent bone biopsies, and final diagnosis of PCM was established according to WHO criteria, and 22 controls. In the present study, all samples were analyzed by flow cytometry (FC) in the side scatter vs CD38 histogram mode, and the CD38-gated plasma cell population was identified. The positive cell ratios of kappa and lambda to plasma cell populations were analyzed. PCM cells were distinguished from normal plasma cells by a cut-off level between 0.80 and 3.3, a sensitivity of 90.3 percent, and a specificity of 81.1 percent. Two-color FC analysis is simple to perform, inexpensive, and clinically relevant data are obtained soon after completion of the FC measurements. It could be one of the helpful tools in the diagnosis of PCM. The correct diagnosis of PCM can be achieved more simply, efficiently, and rapidly by combining this method.

Polychlorinated biphenyls (PCBs) decrease the placental syncytiotrophoblast volume and increase Placental Growth Factor (PlGF) in the placenta of normal pregnancy.

INTRODUCTION: Polychlorinated biphenyls (PCBs) are a class of biologically active, highly stable compounds. Exposure risks include consumption of fatty fish, meat, dairy products and human breast milk, as well as environmental and occupational settings. Numerous reports have described PCB-dependent adverse effects on human fetal growth, including increased risk for IUGR, changes in endocrine function and hormone metabolism, and immunosuppressive and neurological deficits. Here we test the prediction that in utero PCB exposure adversely effects placental morphology, potentially leading to placental insufficiency en route to fetal growth restriction. METHODS: PCB homologs (10) were measured in the maternal and fetal blood of a small cohort of normotensive pregnancies (22) by gas chromatography-mass spectrometry. PCB levels were compared with angiogenesis associated proteins Placental Growth Factor (PlGF) and sFlt-1, determined by ELISA, and the total estimated syncytiotrophoblast (ST) volume. RESULTS: Significant associations between PCB exposure and both PlGF and ST volume were identified. DISCUSSION: PCB effects on placenta morphology and predicted function are discussed. CONCLUSION: These results demonstrate that the human placenta, including ST, is a target of PCB toxicity, and that current environmental PCB exposure levels are a risk to reproductive health.

Medial femoral head border is a reliable and reproducible reference for axis determination for femoral component of unicompartmental knee arthroplasty.

PURPOSE: The femoral component should be implanted parallel to the mechanical axis in unicompartmental knee arthroplasty. It was hypothesised that a line between medial femoral condyle centres and medial border of femoral head will be parallel to the mechanical axis; this study set out to examine this hypothesis. METHODS: One hundred X-rays in fifty patients were included for this study. Long-leg standing X-rays including hip and ankle with patellae facing forwards were obtained. On these films, we measured the angle, α, between mechanical axis and the line between the femoral head centre and knee centre (medial mechanical axis), and the angle, ß, between the medial mechanical axis and a line between medial femoral condyle and femoral head centre. RESULTS: The average value of α was 0.1 ± 0.5° and the average value of ß 3.0° ± 0.3°. These data indicate that mechanical axis and medial mechanical axis are virtually parallel to each other. CONCLUSION: As medial femoral head border is easily identified fluoroscopically, it is a reliable landmark for orientating the femoral component of medial UKA.

Co-expression of multiple cytokines and their receptors in primary clear cell sarcoma of the pubic bone with features of a small round cell tumor.

We present the case of an 81-year-old man with primary clear cell sarcoma (CCS) of the pubic bone with an associated aggressive clinical course. The patient's laboratory tests showed marked leukocytosis, elevated levels of C-reactive protein and multiple cytokines, including interleukin-6 (IL-6) and granulocyte colony-stimulating factor (G-CSF). Histological examination showed monomorphic small cells predominantly arranged as a diffuse sheet with morphological features of a small round cell tumor (SRCT). Immunohistochemical staining indicated that the tumor cells were positive for HMB45, S100, Melan A, IL-6, IL-6 receptor, G-CSF, and G-CSF receptor and negative for cytokeratin (AE1/AE3) and epithelial membrane antigen. To the best of our knowledge, this is the first case report of aggressive primary CCS of the pubic bone with features of SRCT showing the production and co-expression of multiple cytokines and their receptors. Thus, we suggest that proliferation through an IL-6- and G-CSF-associated autocrine mechanism may play an important role in the aggressive clinical course and poor prognosis of some CCSs showing features of SRCT.

Possible involvement of histone acetylation in the development of emotional resistance to stress stimuli in mice.

Recent reports have implied that aberrant biochemical processes in the brain frequently accompany subtle shifts in the cellular epigenetic profile that might underlie the pathogenic progression of psychiatric disorders. Furthermore, certain antidepressants or mood stabilizers have been reported to have the ability to modulate epigenetic parameters. We previously reported that pretreatment of mice with 5-HT(1A) receptor agonists 24 h before testing suppressed the decrease in emotional behaviors induced by exposure to acute restraint stress. Based on this finding, the aim of the present study was to examine the association between the development of emotional resistance to stress stimuli and the modulation of an epigenetic parameter, particularly histone acetylation. We found that acetylated histone H3 was increased in the hippocampus of mice that had developed resistance to emotional stress by pretreatment with flesinoxan (1 mg/kg, i.p.) 24 h before testing. On the other hand, pretreatment with benzodiazepine anxiolytic diazepam (1 mg/kg, i.p.) did not have similar effects. Interestingly, similar to flesinoxan, the histone deacetylase inhibitor trichostatin A also protected against the emotional changes induced by acute restraint stress, as well as histone H3 acetylation. The present findings suggest that the epigenetic mechanisms of gene regulation may play an important role in the development of emotional resistance to stress stimuli.

Rapidly progressive fatal hemorrhagic pneumonia caused by Stenotrophomonas maltophilia in hematologic malignancy.

BACKGROUND: Pneumonia caused by Stenotrophomonas maltophilia is rare, but can be lethal in severely immunocompromised patients. However, its clinical course remains unclear. PATIENTS AND METHODS: Patients with pneumonia caused by S. maltophilia in Toranomon Hospital (890 beds, Tokyo, Japan) were reviewed retrospectively between April 2006 and March 2010. RESULTS: During the study period, 10 cases of S. maltophilia pneumonia were identified. Seven patients had acute myeloid leukemia, 2 had myelodysplastic syndrome, and 1 had malignant lymphoma. All patients developed symptoms after allogeneic hematopoietic stem cell transplantation (HSCT). Five patients received first cord blood transplantation (CBT), 4 patients received second CBT, and 1 patient received first peripheral blood stem cell transplantation (PBSCT). The overall incidence of S. maltophilia pneumonia among 508 patients who received HSCT during the period was 2.0%. The incidence was 0% (0/95) in patients after bone marrow transplantation, 0.8% (1/133) after PBSCT, and 3.2% (9/279) after CBT. Pneumonia developed a median of 13.5 days (range, 6-40) after transplantation. At onset, the median white blood cell count was 10/µL (range, 10-1900), and the median neutrophil count was 0/µL (range, 0-1720). In all patients, S. maltophilia bacteremia developed with bloody sputum or hemoptysis. The 28-day mortality rate was 100%; the median survival after onset of pneumonia was 2 days (range, 1-10). CONCLUSIONS: Hemorrhagic S. maltophilia pneumonia rapidly progresses and is fatal in patients with hematologic malignancy. Attention should be particularly paid to the neutropenic phase early after HSCT or prolonged neutropenia due to engraftment failure. A prompt trimethoprim-sulfamethoxazole-based multidrug combination regimen should be considered to rescue suspected cases of S. maltophilia pneumonia in these severely immunosuppressed patients.

Relationship of intake of plant-based foods with 6-n-propylthiouracil sensitivity and food neophobia in Japanese preschool children.

BACKGROUND/OBJECTIVES: Diets rich in plant-based foods such as vegetables, fruits and soy foods have been suggested to have beneficial effects on health. However, phytochemicals contained in plant-based foods are generally bitter and acrid. We investigated whether intake of vegetables, fruits and soy foods is associated with sensitivity to bitterness and reluctance to eat new foods (food neophobia) in Japanese preschool children. SUBJECTS/METHODS: Subjects of this cross-sectional study were healthy Japanese, 167 boys and 156 girls, aged 4-6 years. Intake of vegetables, fruits and soy foods was estimated from 3-day dietary records. Subjects were classified as either tasters or non-tasters of 6-n-propylthiouracil (PROP) based on their ability to taste 0.56 mmol/l PROP. Information on each child's age, height, weight, food neophobia status and food variety, as well as maternal diet and parental control over the child's eating, was obtained by a parent-administered questionnaire. Food neophobia was assessed using the Child Food Neophobia Scale (CFNS). RESULTS: A high intake of vegetables was significantly associated with a low CFNS score in boys after controlling for covariates (P=0.0008). Among the boys, soy food intake was significantly higher in PROP non-tasters than in tasters, except those with low CFNS scores (P=0.0019). High intake of soy foods was significantly associated with a low neophobia score in PROP tasters but not in non-tasters (P=0.0024). CONCLUSIONS: These data suggest that sensitivity to bitter taste and food neophobia may influence the consumption of vegetables and soy foods among Japanese preschool boys.

Plasma insulin-like peptide 3 and testosterone concentrations in male dogs: changes with age and effects of cryptorchidism.

The objectives were to: (1) develop a time-resolved fluorescence immunoassay (TRFIA) to measure insulin-like peptide 3 (INSL3) in canine plasma; (2) investigate changes of plasma concentrations of INSL3 and testosterone with age in normal male dogs; and (3) compare hormonal concentrations among cryptorchid, normal, and castrated dogs to evaluate endocrine function of the Leydig cell component in retained testes. Blood samples were taken from normal male dogs from prepubertal age to advanced age (4 mo to 14 y, n = 89), and from unilateral cryptorchid (n = 31), bilateral cryptorchid (n = 7), and castrated dogs (n = 3). Canine plasma INSL3 was measured with a newly developed TRFIA. The minimum detection limit of the INSL3 assay was 0.02 ng/ml and the detection range was 0.02 to 20 ng/ml. Plasma INSL3 concentrations increased (P < 0.05) from prepubertal age (4-6 mo) to pubertal age (6-12 mo), and then declined (P < 0.05) from pubertal age to post-pubertal age (1-5 y), reaching a plateau. Plasma testosterone concentrations increased (P < 0.0001) dramatically from prepubertal to pubertal ages, and then seemed to plateau. Concentrations of both INSL3 and testosterone were lower (P < 0.0001 for each) in bilateral cryptorchid dogs than in normal and unilateral cryptorchid dogs. The INSL3 (range: 0.05-0.43 ng/ml) and testosterone (range: 0.10-0.94 ng/ml) concentrations were readily detected in bilateral cryptorchids, but not in castrated dogs (INSL3 < 0.02 ng/ml; testosterone < 0.04 ng/ml). In conclusion, plasma INSL3 concentrations in male dogs measured by a newly developed TRFIA had a transient surge at a pubertal age, whereas testosterone did not. Lower plasma concentrations of INSL3 and testosterone in bilateral cryptorchid dogs suggest impaired endocrine functions of Leydig cell component in paired retained testes. Therefore, peripheral plasma INSL3 and testosterone concentrations have potential diagnostic value in predicting the presence of bilaterally retained testes in male dogs.