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We herein present an extremely rare case of primitive neuroectodermal tumor originating in the penis. A 16-year-old male adolescent presented with painful penile swelling. Pathological, immunohistochemical and cytogenetical examinations of the specimens obtained from total penectomy confirmed the diagnosis of primitive neuroectodermal tumor. After total penectomy, the patient received adjuvant chemotherapy with ifosfamide-based regimen for 48 weeks. As a series of therapies, the patient underwent penile reconstruction surgery after completing adjuvant chemotherapy. The patient has not shown any evidence of recurrence for the 7 years after penile reconstruction surgery, and voiding function is completely normal. A favorable outcome was observed by multimodal therapy including aggressive resection for local control, intensive adjuvant chemotherapy, and penile reconstruction with cosmetic and functional success. Similar therapeutic approaches might be selected for children with primary malignant tumors of the penis., (© 2013 The Japanese Urological Association.)

A novel wavelength combiner using non-uniform refractive index distribution within a multimode interference device is proposed and simulated. The refractive index step creates separate localized modes with different effective refractive indices and two modes are strongly excited which form the basis of an interferometer. We applied the concept to 1.30/1.31 μm and 1.31/1.55 μm wavelength combiners on an InP substrate. The lengths of the devices are 1272 μm and 484 μm with simulated insertion losses of 0.6 dB and 0.67 dB respectively.

In this paper, we examine the performance of several modulation formats in more than four dimensions for coherent optical communications systems. We compare two high-dimensional modulation design methodologies based on spherical cutting of lattices and block coding of a 'base constellation' of binary phase shift keying (BPSK) on each dimension. The performances of modulation formats generated with these methodologies is analyzed in the asymptotic signal-to-noise ratio regime and for an additive white Gaussian noise (AWGN) channel. We then study the application of both types of high-dimensional modulation formats to standard single-mode fiber (SSMF) transmission systems. For modulation with spectral efficiencies comparable to dual-polarization (DP-) BPSK, polarization-switched quaternary phase shift keying (PS-QPSK) and DP-QPSK, we demonstrate SNR gains of up to 3 dB, 0.9 dB and 1 dB respectively, at a BER of 10(-3).

Regulation via interspecific communication is an important for the maintenance of many mutualisms. However, mechanisms underlying the evolution of partner communication are poorly understood for many mutualisms. Here we show, in an ant-lycaenid butterfly mutualism, that attendant ants selectively learn to recognize and interact cooperatively with a partner. Workers of the ant Pristomyrmex punctatus learn to associate cuticular hydrocarbons of mutualistic Narathura japonica caterpillars with food rewards and, as a result, are more likely to tend the caterpillars. However, the workers do not learn to associate the cuticular hydrocarbons of caterpillars of a non-ant-associated lycaenid, Lycaena phlaeas, with artificial food rewards. Chemical analysis revealed cuticular hydrocarbon profiles of the mutualistic caterpillars were complex compared with those of non-ant-associated caterpillars. Our results suggest that partner-recognition based on partner-specific chemical signals and cognitive abilities of workers are important mechanisms underlying the evolution and maintenance of mutualism with ants.

GATA-binding protein 2 (GATA2) and LIM domain only 2 (Lmo2) form common transcription complexes during hematopoietic differentiation. Here we show that these two transcription factors also play a key role in endothelial cells (EC) and lymphatic EC (LEC) function. Primary EC and tumor-associated blood vessels expressed GATA2 and Lmo2. VEGF-induced sprouting angiogenesis in both differentiating embryonic stem cells (embryoid bodies) and primary EC increased GATA2 and Lmo2 levels. Conversely, silencing of GATA2 and Lmo2 expression in primary EC inhibited VEGF-induced angiogenic activity, including EC migration and sprouting in vitro, two key steps of angiogenesis in vivo. This inhibition of EC function was associated with downregulated expression of neuropilin-2 (NRP2), a co-receptor of VEGFRs for VEGF, at the protein, mRNA and promoter levels. NRP2 overexpression partially rescued the impaired angiogenic sprouting in the GATA2/Lmo2 knockdown EC, confirming that GATA2 and Lmo2 mediated EC function, at least in part, by directly regulating NRP2 gene expression. Furthermore, it was found that primary LEC expressed GATA2 and Lmo2 as well. Silencing of GATA2 and Lmo2 expression in LEC inhibited VEGF-induced LEC sprouting, also in a NRP2-dependent manner. In conclusion, our results demonstrate that GATA2 and Lmo2 cooperatively regulate VEGF-induced angiogenesis and lymphangiogenesis via NRP2.

Epoxyeicosatrienoic acids (EETs), lipid mediators produced by cytochrome P450 epoxygenases, regulate inflammation, angiogenesis, and vascular tone. Despite pleiotropic effects on cells, the role of these epoxyeicosanoids in normal organ and tissue regeneration remains unknown. EETs are produced predominantly in the endothelium. Normal organ and tissue regeneration require an active paracrine role of the microvascular endothelium, which in turn depends on angiogenic growth factors. Thus, we hypothesize that endothelial cells stimulate organ and tissue regeneration via production of bioactive EETs. To determine whether endothelial-derived EETs affect physiologic tissue growth in vivo, we used genetic and pharmacological tools to manipulate endogenous EET levels. We show that endothelial-derived EETs play a critical role in accelerating tissue growth in vivo, including liver regeneration, kidney compensatory growth, lung compensatory growth, wound healing, corneal neovascularization, and retinal vascularization. Administration of synthetic EETs recapitulated these results, whereas lowering EET levels, either genetically or pharmacologically, delayed tissue regeneration, demonstrating that pharmacological modulation of EETs can affect normal organ and tissue growth. We also show that soluble epoxide hydrolase inhibitors, which elevate endogenous EET levels, promote liver and lung regeneration. Thus, our observations indicate a central role for EETs in organ and tissue regeneration and their contribution to tissue homeostasis.

Glioblastomas are very difficult tumors to treat because they are highly invasive and disseminate within the normal brain, resulting in newly growing tumors. We have identified netrin-1 as a molecule that promotes glioblastoma invasiveness. As evidence, netrin-1 stimulates glioblastoma cell invasion directly through Matrigel-coated transwells, promotes tumor cell sprouting and enhances metastasis to lymph nodes in vivo. Furthermore, netrin-1 regulates angiogenesis as shown in specific angiogenesis assays such as enhanced capillary endothelial cells (EC) sprouting and by increased EC infiltration into Matrigel plugs in vivo, as does VEGF-A. This netrin-1 signaling pathway in glioblastoma cells includes activation of RhoA and cyclic AMP response element-binding protein (CREB). A novel finding is that netrin-1-induced glioblastoma invasiveness and angiogenesis are mediated by activated cathepsin B (CatB), a cysteine protease that translocates to the cell surface as an active enzyme and co-localizes with cell surface annexin A2 (ANXA2). The specific CatB inhibitor CA-074Me inhibits netrin-1-induced cell invasion, sprouting, and Matrigel plug angiogenesis. Silencing of CREB suppresses netrin-1-induced glioblastoma cell invasion, sprouting, and CatB expression. It is concluded that netrin-1 plays an important dual role in glioblastoma progression by promoting both glioblastoma cell invasiveness and angiogenesis in a RhoA-, CREB-, and CatB-dependent manner. Targeting netrin-1 pathways may be a promising strategy for brain cancer therapy.

A novel polarization splitter on an InP substrate utilizing an MMI coupler loaded with a dielectric and gold layer pad is proposed and simulated. A tilted joint is used for adjusting the phases of TE and TM modes. The MMI section is less than 540 μm. Simulations show that the device has a polarization extinction ratio over 23 dB and an insertion loss below 0.7 dB over the entire C-band for both TE and TM polarizations. The device design was optimized to maximize the wavelength range and tolerance for manufacturing variations.

Unlabelled: What's known on the subject? and What does the study add? Radical cystectomy remains associated with comparatively high perioperative morbidity and mortality, despite improvements in surgical techniques and perioperative care. At present, most studies on the complications associated with open radical cystectomy were derived from Western academic high-volume centres, and data from Japan and other Asian countries were very limited. Using the modified Clavien grading system and 11 category grouping reported from MSKCC, we observed that 68% of patients experienced at least one complication within 90 days of surgery, and 17% of patients experienced major complications (90-day mortality rate = 2%), which were compatible with reports from Western high-volume centres. As far as we know, our report is the largest one regarding perioperative morbidity and mortality in Asian patients who underwent radical cystectomy., Objective: To determine the type, incidence and severity of 90-day morbidity after radical cystectomy in our institution and our affiliated hospitals in accordance with a standard reporting methodology. At present, most studies on complications associated with open radical cystectomy are derived from Western academic high-volume centres and data from Japan and other Asian countries remain very limited., Patients and Methods: The study comprised a retrospective multi-institutional study. The records were reviewed of 928 patients who underwent open radical cystectomy between 1997 and 2010. All complications within 90 days of surgery were categorized into 11 specific categories and graded in accordance with the modified Clavien system. Multivariate regression models were used to determine predictors of complications., Results: At least one complication was observed in 635 (68%) patients and a major (grade 3-5) complication was observed in 156 (17%) patients. The most common complication categories were infectious (30%), gastrointestinal (26%), wound-related (21%) and genitourinary (15%). The 30-day mortality rate was 0.8% and the 90-day mortality rate was 2%. A multivariate regression model showed that previous cardiovascular comorbidity and type of urinary diversion (i.e. ileal conduit or neobladder) were significant factors for any and major complications., Conclusions: Surgical complication-related radical cystectomy is significant and both previous cardiovascular comorbidity and the type of urinary diversion were found to be significant factors for any and major complications. The 90-day mortality rate was 2%, which is compatible with reports from Western high-volume centres., (© 2012 THE AUTHORS. BJU INTERNATIONAL © 2012 BJU INTERNATIONAL.)

Fiber nonlinearity has become a major limiting factor to realize ultra-high-speed optical communications. We propose a fractionally-spaced equalizer which exploits a trained high-order statistics to deal with data-pattern dependent nonlinear impairments in fiber-optic communications. The computer simulation reveals that the proposed 3-tap equalizer improves Q-factor by more than 2 dB for long-haul transmissions of 5,230 km distance and 40 Gbps data rate. We also demonstrate that the joint use of a digital backpropagation (DBP) and the proposed equalizer offers an additional 1-2 dB performance improvement due to the channel shortening gain. A performance in high-speed transmissions of 100 Gbps and beyond is evaluated as well.

A mode-evolution-based polarization rotator-splitter built on InP substrate is proposed by combining a mode converter and an adiabatic asymmetric Y-coupler. The mode converter, consisting of a bi-level taper and a width taper, effectively converts the fundamental TM mode into the second order TE mode without changing the polarization of the fundamental TE mode. The following adiabatic asymmetric Y-coupler splits the fundamental and the second order TE modes and also converts the second order TE mode into the fundamental TE mode. A shallow etched structure is proposed for the width taper to enhance the polarization conversion efficiency. The device has a total length of 1350 µm, a polarization extinction ratio over 25 dB and an insertion loss below 0.5 dB both for TE and TM modes, over the wavelength range from 1528 to 1612 nm covering all C + L band. Because the device is designed based on mode evolution principle, it has a large fabrication tolerance. The insertion loss remains below 1 dB and the polarization extinction ratio remains over 17 dB with respect to a width variation of +/- 0.12 µm at the wavelength of 1570 nm, or +/- 0.08 µm over the entire C + L band., (© 2012 Optical Society of America)

Tumor endothelial cells (TECs) are therapeutic targets in anti-angiogenic therapy. Contrary to the traditional assumption, TECs can be genetically abnormal and might also acquire drug resistance. In this study, mouse TECs and normal ECs were isolated to investigate the drug resistance of TECs and the mechanism by which it is acquired. TECs were more resistant to paclitaxel with the up-regulation of multidrug resistance (MDR) 1 mRNA, which encodes the P-glycoprotein, compared with normal ECs. Normal human microvascular ECs were cultured in tumor-conditioned medium (CM) and became more resistant to paclitaxel through MDR1 mRNA up-regulation and nuclear translocation of Y-box-binding protein 1, which is an MDR1 transcription factor. Vascular endothelial growth factor (VEGF) receptor 2 (VEGFR2) and Akt were activated in human microvascular ECs by tumor CM. We observed that tumor CM contained a significantly high level of VEGF. A VEGFR kinase inhibitor, Ki8751, and a phosphatidylinositol 3-kinase-Akt inhibitor, LY294002, blocked tumor CM-induced MDR1 up-regulation. MDR1 up-regulation, via the VEGF-VEGFR pathway in the tumor microenvironment, is one of the mechanisms of drug resistance acquired by TECs. We observed that VEGF secreted from tumors up-regulated MDR1 through the activation of VEGFR2 and Akt. This process is a novel mechanism of the acquisition of drug resistance by TECs in the tumor microenvironment., (Copyright © 2012 American Society for Investigative Pathology. Published by Elsevier Inc. All rights reserved.)

The results of biocontrol with entomopathogens in termites have been discouraging because of the strong social hygiene behavior for removing pathogens from termite colonies. However, the mechanism of pathogen detection is still unclear. For the successful application of biopesticides to termites in nature, it would be beneficial to identify substances that could disrupt the termite's ability to perceive pathogens. We hypothesized that termites can perceive pathogens and this ability plays an important role in effective hygiene behavior. In this study, pathogen-detection in the subterranean termite Coptotermes formosanus was investigated. We performed quantitative assays on conidia removal by grooming behavior using epifluoresence microscopy and Y-maze tests to examine the perception of fungal odor by termites. Three species each of high- and low-virulence entomopathogenic fungi were used in each test. The results demonstrated that termites removed conidia more effectively from a nestmate's cuticle if its odor elicited stronger aversion. Highly virulent pathogens showed higher attachment rates to termite surfaces and their odors were more strongly avoided than those of low-virulence isolates in the same species. Moreover, termites appeared to groom each other more persistently when they had more conidia on their bodies. In brief, insect perception of pathogen-related odor seems to play a role in the mechanism of their hygiene behavior.

Termites often eliminate pathogens directly through mutual grooming, and are thereby prevent infections from entomopathogenic fungi. Our previous study confirmed that the antennae of Coptotermesformosanus sensitively responded to the musty odor of entomopathogenic fungi. However, it is unclear if this odor has any effect on termite behavior. The purpose of this study was to clarify the effects of fungal odor on termite behavior, especially on conidia removal. The musty odor was prepared as an aqueous solution by immersing conidia in distilled water. When untreated termites were mixed with fungal-odor-treated termites at a ratio of 4:1, mutual grooming and attack of treated termites were frequently observed. This indicated that the fungal odor triggered these behavioral responses. While some components of the fungal odor were found in all of the entomopathogenic fungi tested, the odor profiles differed among the isolates., (Copyright © 2011 Elsevier Inc. All rights reserved.)

The studies of pathogen-prevention behaviors of termites have focused on hygiene behavior directed only against highly virulent pathogens. Therefore, we compared behavioral changes in the subterranean termite Coptotermes formosanus following contact with entomopathogenic fungi with different levels of virulence. The fungal virulence was inferred from the daily mortality and the LD50 value in previous data. When untreated termites were allowed to contact their fungus-inoculated nestmates, mutual grooming was frequent during 30 min after inoculation. The inoculated termites were often attacked and eaten by their uninoculated nestmates, and then buried after death. Notably, there was no influence of fungal virulence on these pathogen-prevention behaviors. However, the fungal isolates and genera affected not only the frequency of the behaviors but also the horizontal transmission pattern, the number of dead individuals and the survival period before the first death following infection., (Copyright © 2011 Elsevier Inc. All rights reserved.)

We identified the soldier-specific compounds in the Japanese subterranean termite, Reticulitermes speratus, to clarify their ethological roles. Silica gel column chromatography separated one major soldier-specific compound in the hexane fraction accounting for 70-80% of the total amount of the fraction, while cuticular hydrocarbons constituted the rest. We identified the compound as β-selinene by gas chromatography-mass spectrometry (GC-MS) and nuclear magnetic resonance (NMR) spectroscopy. Comparative GC analyses of the major exocrine glands detected the compound in the soldier's frontal gland. Both soldiers and workers made aggregation to the hexane fraction, as well as to the crushed heads and head extract of the soldiers. They did not aggregate to cuticular hydrocarbons, making it likely that β-selinene was the aggregation pheromone in this species. The opportunistic predator of this termite, Lasius japonicus, was also attracted to the compounds. The ant workers, therefore, would use the termite aggregation pheromone as a kairomone for hunting them.

Adrenomedullin (AM) is a multifunctional 52-amino acid peptide. AM has several effects and acts as a growth factor in several types of cancer cells. Our previous study revealed that an AM antagonist (AMA) suppressed the growth of pancreatic tumors in mice, although its mechanism was not elucidated. In this study, we constructed an AMA expression vector and used it to treat renal cell carcinoma (RCC) in mice. This AMA expression vector significantly reduced tumor growth in mice. In addition, microvessel density was decreased in AMA-treated tumors. To analyze the effect of AMA on tumor angiogenesis in this model, tumor endothelial cells (TECs) were isolated from RCC xenografts. TEC proliferation was stimulated by AM and it was inhibited by AMA significantly. AM induced migration of TECs and it was also blocked by AMA. However, normal ECs (NECs) were not affected by either AM or AMA. These results demonstrate that AMA has inhibitory effects on TECs specifically, not on NEC, thereby inhibiting tumor angiogenesis. Furthermore, we showed that vascular endothelial growth factor-induced mobilization of endothelial progenitor cell (EPC) into circulation was inhibited by AMA. These results suggest that AMA can be considered a good anti-angiogenic reagent that selectively targets TECs and EPC in renal cancer.

Tumor angiogenesis is necessary for solid tumor progression and metastasis. Increasing evidence indicates that tumor endothelial cells (TECs) are more relevant to the study of tumor angiogenesis than normal endothelial cells (NECs) because their morphologies and gene expression are different from NECs. However, it is challenging to isolate and culture large numbers of pure ECs from tumor tissue since the percentage of ECs is only about 1-2% and tumor cells and fibroblasts easily overgrow them. In addition, there has been concern that isolated TECs may lose their special phenotype once they are dissociated from tumor cells. In this study, we have successfully purified murine TECs from four different human tumor xenografts and NECs from murine dermal tissue. Isolated ECs expressed endothelial markers, such as CD31, VE-cadherin (CD144), and endoglin (CD105), for more than 3 months after isolation. TECs maintained tumor endothelial-specific markers, such as tumor endothelial marker 8 (TEM8) and aminopeptidase N (APN), as in tumor blood vessels in vivo. In addition, TECs were more proliferative and motile than NECs. TECs showed a higher response to VEGF and higher expression of VEGF receptors-1 and -2 than NECs did. Stem cell antigen-1 was up-regulated in all four TECs, suggesting that they have a kind of stemness. Cultured TECs maintain distinct biological differences from NECs as in vivo. In conclusion, it was suggested that TECs are relevant material for tumor angiogenesis research., (2010 Elsevier Inc. All rights reserved.)

Objectives: To determine the pathological features and clinical course of intravesical recurrence after nephroureterectomy (NU) for upper urinary tract (UUT) cancer., Patients and Methods: Among 325 patients undergoing NU with bladder cuff excision for UUT cancer, in this retrospective multi-institutional study we evaluated 113 who developed bladder tumour after NU. Excluding patients with (i) perioperative systemic chemotherapy or radiotherapy for UUT cancer; (ii) a history of previous or synchronous bladder cancer at the time of NU; (iii) distant metastasis at the time of NU; (iv) a follow-up of <1 year after the initial bladder cancer recurrence; or (v) missing data, 74 patients were included in this study. We compared the pathology between UUT cancer and the first bladder cancer recurrence, using Fisher's exact test. Further intravesical recurrence and bladder cancer progression was analysed using the Kaplan-Meier method, with the log-rank test used to assess significance. A Cox proportional hazard model was used for multivariate analysis., Results: The grade of the first bladder cancer recurrence strongly correlated with that of the UUT tumour (P < 0.001) and the carcinoma in situ (CIS) lesion with the first bladder cancer recurrence correlated with high grade (grade 3) UUT tumour (P < 0.001). In all, 56 of the assessable 70 patients further developed intravesical recurrence at a median interval of 7 months after the first bladder cancer recurrence. There were no clinicopathological factors that predicted the second recurrence. Progression occurred in 14 patients, at a median interval of 25 months. A CIS lesion with the first bladder cancer recurrence was a risk factor for progression on multivariate analysis., Conclusions: A large proportion of the patients who developed bladder tumour after NU had further intravesical recurrence, which indicated its refractory nature. Especially when a CIS lesion is detected in the initial intravesical recurrence, a careful follow-up is mandatory to detect bladder cancer progression., (© 2009 THE AUTHORS. JOURNAL COMPILATION © 2009 BJU INTERNATIONAL.)

Our experience in the design of an ultra-high speed image sensor targeting the theoretical maximum frame rate is summarized. The imager is the backside illuminated in situ storage image sensor (BSI ISIS). It is confirmed that the critical factor limiting the highest frame rate is the signal electron transit time from the generation layer at the back side of each pixel to the input gate to the in situ storage area on the front side. The theoretical maximum frame rate is estimated at 100 Mega-frames per second (Mfps) by transient simulation study. The sensor has a spatial resolution of 140,800 pixels with 126 linear storage elements installed in each pixel. The very high sensitivity is ensured by application of backside illumination technology and cooling. The ultra-high frame rate is achieved by the in situ storage image sensor (ISIS) structure on the front side. In this paper, we summarize technologies developed to achieve the theoretical maximum frame rate, including: (1) a special p-well design by triple injections to generate a smooth electric field backside towards the collection gate on the front side, resulting in much shorter electron transit time; (2) design technique to reduce RC delay by employing an extra metal layer exclusively to electrodes responsible for ultra-high speed image capturing; (3) a CCD specific complementary on-chip inductance minimization technique with a couple of stacked differential bus lines.

Averaging of accumulated data is a standard technique applied to processing data with low signal-to-noise ratios (SNR), such as image signals captured in ultra-high-speed imaging. The authors propose an architecture layout of an ultra-high-speed image sensor capable of on-chip signal accumulation. The very high frame rate is enabled by employing an image sensor structure with a multi-folded CCD in each pixel, which serves as an in situ image signal storage. The signal accumulation function is achieved by direct connection of the first and the last storage elements of the in situ storage CCD. It has been thought that the multi-folding is achievable only by driving electrodes with complicated and impractical layouts. Simple configurations of the driving electrodes to overcome the difficulty are presented for two-phase and four-phase transfer CCD systems. The in situ storage image sensor with the signal accumulation function is named Image Signal Accumulation Sensor (ISAS).

Tumor blood vessels are thought to contain genetically normal and stable endothelial cells (ECs), unlike tumor cells, which typically display genetic instability. Yet, chromosomal aberration in human tumor-associated ECs (hTECs) in carcinoma has not yet been investigated. Here we isolated TECs from 20 human renal cell carcinomas and analyzed their cytogenetic abnormalities. The degree of aneuploidy was analyzed by fluorescence in situ hybridization using chromosome 7 and chromosome 8 DNA probes in isolated hTECs. In human renal cell carcinomas, 22-58% (median, 33%) of uncultured hTECs were aneuploid, whereas normal ECs were diploid. The mechanisms governing TEC aneuploidy were then studied using mouse TECs (mTECs) isolated from xenografts of human epithelial tumors. To investigate the contribution of progenitor cells to aneuploidy in mTECs, CD133(+) and CD133(-) mTECs were compared for aneuploidy. CD133(+) mTECs showed aneuploidy more frequently than CD133(-) mTECs. This is the first report showing cytogenetic abnormality of hTECs in carcinoma, contrary to traditional belief. Cytogenetic alterations in tumor vessels of carcinoma therefore can occur and may play a significant role in modifying tumor- stromal interactions.

The exploitation of parental care is common in avian and insect 'cuckoos' and these species engage in a coevolutionary arms race. Caterpillars of the lycaenid butterfly Niphanda fusca develop as parasites inside the nests of host ants (Camponotus japonicus) where they grow by feeding on the worker trophallaxis. We hypothesized that N. fusca caterpillars chemically mimic host larvae, or some particular castes of the host ant, so that the caterpillars are accepted and cared for by the host workers. Behaviourally, it was observed that the host workers enthusiastically tended glass dummies coated with the cuticular chemicals of larvae or males and those of N. fusca caterpillars living together. Cuticular chemical analyses revealed that N. fusca caterpillars grown in a host ant nest acquired a colony-specific blend of cuticular hydrocarbons (CHCs). Furthermore, the CHC profiles of the N. fusca caterpillars were particularly close to those of the males rather than those of the host larvae and the others. We suggest that N. fusca caterpillars exploit worker care by matching their cuticular profile to that of the host males, since the males are fed by trophallaxis with workers in their natal nests for approximately ten months.

Background: Ants of the genus Lasius are ecologically important and an important system for evolutionary research. Progress in evolutionary research has been hindered by the lack of a well-founded phylogeny of the subgenera, with three previous attempts disagreeing. Here we employed two mitochondrial genes (cytochrome c oxidase subunit I, 16S ribosomal RNA), comprising 1,265 bp, together with 64 morphological characters, to recover the phylogeny of Lasius by Bayesian and Maximum Parsimony inference after exploration of potential causes of phylogenetic distortion. We use the resulting framework to infer evolutionary pathways for social parasitism and fungiculture., Results: We recovered two well supported major lineages. One includes Acanthomyops, Austrolasius, Chthonolasius, and Lasius pallitarsis, which we confirm to represent a seventh subgenus, the other clade contains Dendrolasius, and Lasius sensu stricto. The subgenus Cautolasius, displaying neither social parasitism nor fungiculture, probably belongs to the second clade, but its phylogenetic position is not resolved at the cutoff values of node support we apply. Possible causes for previous problems with reconstructing the Lasius phylogeny include use of other reconstruction techniques, possibly more prone to instabilities in some instances, and the inclusion of phylogenetically distorting characters., Conclusion: By establishing an updated phylogenetic framework, our study provides the basis for a later formal taxonomic revision of subgenera and for studying the evolution of various ecologically and sociobiologically relevant traits of Lasius, although there is need for future studies to include nuclear genes and additional samples from the Nearctic. Both social parasitism and fungiculture evolved twice in Lasius, once in each major lineage, which opens up new opportunities for comparative analyses. The repeated evolution of social parasitism has been established for other groups of ants, though not for temporary social parasitism as found in Lasius. For fungiculture, the independent emergence twice in a monophyletic group marks a novel scenario in ants. We present alternative hypotheses for the evolution of both traits, with one of each involving loss of the trait. Though less likely for both traits than later evolution without reversal, we consider reversal as sufficiently plausible to merit independent testing.

Objectives: We retrospectively analyzed the survival of renal cell carcinoma patients with bone metastases, and identified prognostic factors and a model predictive for survival in these patients., Methods: Fifty patients with renal cell carcinoma with osseous metastases were treated at Hokkaido University Hospital between 1980 and 2004. The relationship between several clinical features and survival was examined univariately. The Cox proportional hazards model was then used to form a multivariate model., Results: The median survival time from the diagnosis of bone metastasis was 12 mo, and overall survival at 2 yr was 37%. Clinical features correlated with longer survival in the multivariate analysis were a long interval (24 mo or more) between the diagnosis of kidney cancer and that of osseous metastasis (hazard ratio [HR]: 2.608; 95% confidence interval [CI], 1.031-6.599) and the absence of extraosseous metastases (HR: 2.523; 95%CI, 1.023-6.220). By combining these two favorable factors, renal cell carcinoma patients with osseous metastases could be categorized into two different groups. The median time to death in 20 patients with zero favorable factors (poor prognosis) was 5 mo. On the other hand, 30 patients had one or two favorable factors (good prognosis); the median survival time in this group was 30 mo. There was a significant difference in survival duration between the two groups (p<0.001)., Conclusions: Two prognostic factors predicting survival were identified and used to categorize renal cell carcinoma patients with bone metastasis into two prognostic groups.

Wax, 85% of which consists of orthorhombic crystals, has been found in the cocoon of the hornet Vespa analis Fabricius by means of high-resolution (13)C solid-state nuclear magnetic resonance (NMR). GC-MS analysis revealed the major components of the wax in the cocoon were linear alkenes and alkanes with a total of 23 or 34 carbon atoms. At 40.7 degrees C a DSC absorption peak and a (13)C NMR chemical shift change were observed and interpreted as the result of a crystal transition from the orthorhombic to rotator phase of the wax molecules. At 55.5 degrees C melting of the wax was observed. The amount of crystalline wax deposition varied with the part of the cocoon-crystalline wax was concentrated in the silk sleeve lining the inner wall of each comb cell but there was very little in the silk cap projecting from the end of each cell. Because the wax components of the larval cocoon were almost identical to those of the larval cuticle, despite a slight difference in the profiles, they might have come from the larval cuticle via direct body contact with the cocoon.

The mechanism by which palatable species take advantage of their similarity in appearance to those that are unpalatable, in order to avoid predation, is called Batesian mimicry. Several arthropods are thought to be Batesian mimics of social insects; however, social insects that are Batesian mimics among themselves are rare. In Malaysia we found a possible Batesian mimic in an arboreal ant species, Camponotus sp., which was exclusively observed on foraging trails of the myrmicine ant Crematogaster inflata. The bright yellow and black colouring pattern, as well as the walking behaviour, were very similar in both species. We observed general interactions between the two species, and tested their palatability and the significance of the remarkably similar visual colour patterns for predator avoidance. Prey offered to C. inflata was also eaten by Camponotus workers in spite of their being attacked by C. inflata, indicating that Camponotus sp. is a commensal of C. inflata. An experiment with chicks as potential predators suggests that Camponotus sp. is palatable whereas C. inflata is unpalatable. After tasting C. inflata, the chicks no longer attacked Camponotus sp., indicating that Camponotus sp. is a Batesian mimic of Crematogaster inflata.

Cuticular hydrocarbons and morphological features were compared among 80 Formica japonica colonies collected in Japan. Although a few morphological differences were found in workers among the colonies, four different types of cuticular hydrocarbon composition were observed. This was supported by a principal component analysis. We further compared the cuticular hydrocarbons among a total of approximately 400 F. japonica colonies, and categorized the hydrocarbon components into four types based on the result of discriminant analyses for the first 80 colonies. Type 1 was observed in colonies mainly collected in southern Honshu, Shikoku, and Kyushu. Types 2, 3, and 4 were from colonies with primary collections in Southern Honshu, central and Pacific coast northern Honshu, and the Sea of Japan coasts of northern Honshu and Hokkaido, respectively. The occurrence of four distinct types of CHC composition suggests that the colonies that produce them are separate species.

Toll-like receptor 2 (TLR2) has been recognized to mediate cell signaling in response to peptidoglycan (PGN), a major cell wall component of Gram-positive bacteria. The mechanism by which TLR2 recognizes PGN is unknown. It is not even clear whether TLR2 directly binds to PGN. In this study, we generated a soluble form of recombinant TLR2 (sTLR2) possessing only its putative extracellular domain by using the baculovirus expression system to examine the direct interaction between sTLR2 and PGN. sTLR2 bound avidly to insoluble PGN (iPGN) from Staphylococcus aureus coated onto microtiter wells in a concentration-dependent manner. In contrast, sTLR2 exhibited a very weak binding to lipopolysaccharide. iPGN cosedimented sTLR2 after the mixture of iPGN and sTLR2 had been incubated and centrifuged. sTLR2 partially attenuated the iPGN-induced NF-kappaB activation in TLR2-transfected HEK 293 cells and the iPGN-induced IL-8 secretion in U937 cells. One of anti-human TLR2 monoclonal antibodies, which blocked iPGN-induced NF-kappaB activation in TLR2-transfected cells, inhibited the binding of sTLR2 to iPGN. In addition, we found that sCD14 interacted with sTLR2 and increased the binding of sTLR2 to iPGN. From these results, we conclude that the extracellular TLR2 domain directly binds to PGN.

Pulmonary surfactant protein A (SP-A) plays an important role in modulation of the innate immune system of the lung. Peptidoglycan (PGN), a cell wall component of Gram-positive bacteria, is known to elicit excessive proinflammatory cytokine production from immune cells. In this study we investigated whether SP-A interacts with PGN and alters PGN-elicited cellular responses. Binding studies demonstrate that PGN is not a ligand for SP-A. However, SP-A significantly reduced PGN-elicited tumor necrosis factor alpha (TNF-alpha) secretion by U937 cells and rat alveolar macrophages. The inhibitory effect on TNF-alpha secretion was dependent upon SP-A concentrations in physiological range. Coincubation of SP-A and PGN with human embryonic kidney 293 cells that had been transiently transfected with the cDNA of Toll-like receptor 2 (TLR2), a cell signaling receptor for PGN, significantly attenuated PGN-induced nuclear factor-kappaB activity. SP-A directly bound to a soluble form of the recombinant extracellular TLR2 domain (sTLR2). Coincubation of sTLR2 with SP-A significantly reduced the binding of sTLR2 to PGN. These results indicate that the direct interaction of SP-A with TLR2 alters PGN-induced cell signaling. We propose that SP-A modulates inflammatory responses against the bacterial components by interactions with pattern-recognition receptors.

The roles of the four domains of annexin IV in binding to phospholipids and glycolipids were assessed by analyzing the binding of a group of mutant annexins IV in which one or more of the four domains was inactivated by replacing a critical amino residue(s) (Asp or Glu) with the neutral residue Ala. The data reveal that individual annexin domains may have characteristic affinities for different lipids. In particular, inactivation of the fourth domain inhibits the binding to phosphatidylserine (PS) and phosphatidylinositol (PI) but not to phosphatidylglycerol (PG), suggesting that this domain specifically can accommodate the larger head groups of PS and PI whereas the other three domains may form more restricted binding pockets. In order to block binding to PG, domain 1, or both domains 2 and 3 must be inactivated in addition to domain 4, suggesting that all four domains may be able to accommodate the headgroup of PG to some extent. Binding to acidic glycolipids (sulfatides) was also sensitive to inactivation of domain 4. However, in the case of sulfatides the nature of the binding reaction is fundamentally different compared with the binding to phospholipids since the interaction with sulfatides was highly sensitive to an increase in ionic strength. The binding to sulfatides may depend therefore on charge-charge interactions whereas the binding to phospholipid may involve a more specific interaction between the lipid headgroup and the protein surface, and/or interaction of the protein with the hydrophobic portion of a lipid bilayer.

Surfactant proteins A and D (SP-A and SP-D) are structurally related members of the collectin family found in the alveolar compartment of the lung. SP-A binds dipalmitoylphosphatidylcholine (DPPC) and galactosylceramide (GalCer), induces liposome aggregation, and regulates the uptake and secretion of surfactant lipids by alveolar type II cells in vitro. SP-D binds phosphatidylinositol (PI) and glucosylceramide. The purpose of this study was to identify a critical stretch of primary sequence in the SP-A region Cys(204)-Phe(228) and the SP-D region Cys(331)-Phe(355) that is involved in protein-specific lipid and type II cell interactions. Chimeras ad1 and ad2 were constructed with rat SP-A/SP-D splice junctions at Cys(218)/Gly(346) and Lys(203)/Cys(331), respectively. Chimera ad1 but not ad2 retained DPPC liposome binding activity. Both chimeras retained significant binding to GalCer liposomes. Chimera ad1 did not bind to PI, whereas chimera ad2 acquired a significant PI binding. Both chimeras failed to induce liposome aggregation and to interact with alveolar type II cells. In addition, monoclonal antibody 1D6 that blocks specific SP-A functions did not recognize either chimera. From these results, we conclude that (1) the SP-A region Leu(219)-Phe(228) is required for liposome aggregation and interaction with alveolar type II cells, (2) the SP-A region Cys(204)-Cys(218) is required for DPPC binding, (3) the SP-D region Cys(331)-Phe(355) is essential for minimal PI binding, and (4) the epitope for mAb 1D6 is located at the region contiguous to the SP-A region Leu(219)-Phe(228).

We examined guanosine triphosphate (GTP)-binding proteins (G-proteins) in synovial tissues obtained from patients with rheumatoid arthritis or osteoarthritis. The results of immunoblot analysis with anti-bovine G-protein betagamma-subunit antibody in the rheumatoid synovial tissue were similar to those in the osteoarthritis synovial tissue. On the other hand, the carboxyl methylation activity on G-protein gamma-subunit in the rheumatoid synovial tissue was enhanced compared with that in the osteoarthritis synovial tissues: Km and Vm values were 2.6 microM and 10 pmol/mg x min, respectively, for the rheumatoid arthritis, and 4.8 microM and 5.6 pmol/mg x min, respectively, for the osteoarthritis. These results suggest that G-protein-linked signal transduction, in reference to carboxyl methylation of the gamma-subunit, is affected in rheumatoid tissues.

The goal of this study was to determine the changes that occur in surfactant-associated proteins in bronchoalveolar lavage fluid (BAL) and serum of patients at risk for ARDS and during the course of ARDS. We found that the concentrations of SP-A and SP-B were low in the BAL of patients at risk for ARDS before the onset of clinically defined lung injury, whereas the concentration of SP-D was normal. In patients with established ARDS, BAL SP-A and SP-B concentrations were low during the entire 14-d observation period, but the median SP-D concentrations remained in the normal range. Immunoreactive SP-A and SP-D were not increased in the serum of patients at risk for ARDS, but both increased after the onset of ARDS to a maximum on Day 3 and remained elevated for as long as 14 d. The BAL SP-A concentrations were significantly lower in at-risk patients who developed ARDS, and no patient with a BAL SP-A concentration greater than 1.2 microg/ml developed ARDS. On Days 1 and 3 of ARDS, the BAL SP-D concentration was significantly lower in patients who died, and the BAL SP-D concentration was significantly related to the PI(O(2))/FI(O(2)) ratio. Thus, surfactant protein abnormalities occur before and after the onset of ARDS, and the responses of SP-A, SP-B, and SP-D differ in important ways. The BAL SP-A and SP-D measurements can be used to classify patients as high or low risk for progression to ARDS and/or death after the onset of ARDS. Strategies to increase these surfactant proteins in the lungs of patients with ARDS could be useful to modify the onset or the course of ARDS.

We have shown previously that surfactant protein A (SP-A) binds to annexin IV in a Ca2+-dependent manner [Sohma, Matsushima, Watanabe, Hattori, Kuroki and Akino (1995) Biochem. J. 312, 175-181]. Annexin IV is a member of the annexin family having four consensus repeats of about 70 amino acids and a unique N-terminal tail. In the present study, the functional site of both annexin IV and SP-A for the Ca2+-dependent binding was investigated using mutant proteins. SP-A bound in a Ca2+-dependent manner to an annexin-IV truncation mutant consisting of the N-terminal domain and the first three domains (T(N-1-2-3)). SP-A also bound to T3-4, but this interaction was not Ca2+-dependent. SP-A bound weakly to the other truncation mutants (T(N-1-2), T(2-3) and T(2-3-4)). Each consensus repeat of annexin IV possesses a conserved acidic amino acid residue (Glu70, Asp142, Glu226 and Asp301) that putatively ligates Ca2+. Using annexin-IV DE mutants in which one, two or three residues out of the four Asp/Glu were altered to Ala by site-directed mutagenesis [Nelson and Creutz (1995) Biochemistry 34, 3121-3132], it was revealed that Ca2+ binding in the third domain is more important than in the other Ca2+-binding sites. SP-A is a member of the animal lectin group homologous with mannose-binding protein A. The substitution of Arg197 of rat SP-A with Asp or Asn eliminated binding to annexin IV, whereas the substitution of Glu195 with Gln was silent. These results suggest that the Ca2+ binding to domain 3 of annexin IV is required for the Ca2+-dependent binding by SP-A and that Arg197 of SP-A is important in this binding.

Pulmonary surfactant protein A (SP-A) and mannose-binding protein A (MBP-A) are collectins in the C-type lectin superfamily. These collectins exhibit unique lipid binding properties. SP-A binds to dipalmitoyl phosphatidylcholine (DPPC) and galactosylceramide (GalCer) and MBP-A binds to phosphatidylinositol (PI). SP-A also interacts with alveolar type II cells. Monoclonal antibodies (mAbs PE10 and PC6) that recognize human SP-A inhibit the interactions of SP-A with lipids and alveolar type II cells. We mapped the epitopes for anti-human SP-A mAbs by a phage display peptide library. Phage selected by mAbs displayed the consensus peptide sequences that are nearly identical to 184TPVNYTNWYRG194 of human SP-A. The synthetic peptide GTPVNYTNWYRG completely blocked the binding of mAbs to human SP-A. Chimeric proteins were generated in which the rat SP-A region Thr174-Gly194 or the human SP-A region Ser174-Gly194 was replaced with the MBP-A region Thr164-Asp184 (rat ama4 or hu ama4, respectively). The mAbs failed to bind hu ama4. Rat ama4 bound to an affinity matrix on mannose-sepharose but lost all of the SP-A functions except carbohydrate binding and Ca2+-independent GalCer binding. Strikingly, the rat ama4 chimera acquired the PI binding property that MBP-A exhibits. This study demonstrates that the amino acid residues 174-194 of SP-A and the corresponding region of MBP-A are critical for SP-A-type II cell interaction and Ca2+-dependent lipid binding of collectins.

Objective: To characterize surfactant protein D (SP-D) isolated from bronchoalveolar lavage fluid (BALF) of healthy horses., Sample Population: BALF from 10 Thoroughbreds (5 males, 5 females; 26 to 40 months old) without history or clinical signs of respiratory tract disease., Procedure: BALF was obtained and centrifuged at 33,000 X g. The supernatant was applied to a mannose-Sepharose 6B affinity column in the presence of calcium, and the bound protein fraction was analyzed by use of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, immunoblot analysis; amino acid composition was determined and partial sequencing was done. Phospholipid binding and liposome aggregation assay were performed, using purified proteins., Results: The protein isolated by use of mannose affinity matrices was SP-D. It bound carbohydrates and phosphatidylinositol, which are the characteristic features of SP-D isolated from other animal species. Amino acid analysis and partial primary sequence of the isolated protein indicated high homology with rat and human SP-D. Furthermore, immunoblot analysis indicated that equine SP-D reacted with human and rat SP-D-specific antibodies., Conclusion and Clinical Relevance: SP-D exists in equine lungs; its measurement may be useful in evaluating equine lung disease.

Objective: To characterize surfactant protein isolated from bronchoalveolar lavage fluids of healthy horses., Animals: 10 Thoroughbreds (5 males, 5 females; 26 to 40 months old) that did not have a history or clinical signs of respiratory tract disease., Procedure: Bronchoalveolar lavage fluid (BALF) was obtained and centrifuged at 33,000 X g. Lipid was removed from precipitated fractions by means of extraction with 1-butanol, and organic solvent-insoluble protein precipitates were dialyzed against Tris buffer. The suspension was centrifuged, and supernatant was placed in a mannose-Sepharose affinity column, with calcium. The bound protein fraction was analyzed by means of sodium dodecyl sulfate-polyacrylamide gel electrophoresis, western immunoblot analysis, and amino acid sequencing. A liposome-aggregation assay was also performed, using purified proteins., Results: Protein isolated by use of mannose-affinity matrices was identified as surfactant protein A (SP-A). It had carbohydrate-binding and phospholipid-aggregation properties characteristic of SP-A isolated from other animal species. The partial primary sequence of the isolated protein had high homology with rat and human SP-A. Furthermore, the equine SP-A reacted with anti-human and anti-rat SP-A specific antibodies., Conclusion: Analysis of these findings indicated the existence of SP-A in pulmonary tissues of horses., Clinical Relevance: Measurement of SP-A concentrations may be useful for clinicians evaluating pulmonary disease of horses.

The purpose of this study was to characterize the toxic effects of lysophosphatidylcholine (lyso-PC) on neonatal lung function. Various doses of lyso-PC (from 0 to 40 mg/kg) were administered to near-term newborn rabbits. Lung-thorax compliance during mechanical ventilation was significantly decreased by doses >/=10 mg/kg, and static lung volumes during deflation were decreased by doses >/=20 mg/kg. Using the same experimental model, we investigated the effects of modified porcine surfactant (Curosurf, 200 mg/kg). Animals exposed to lyso-PC at birth and treated simultaneously with surfactant showed a satisfactory therapeutic response, whereas those treated after 30 min failed to respond. These animals also had a much larger leak of albumin into the air spaces and an elevated minimum surface tension of the lavage fluid in a pulsating bubble surfactometer, suggesting inactivation of the exogenous surfactant. Timing of surfactant administration may thus be essential for the therapeutic effect in this experimental model of acute lung injury.

To elucidate the effect of a modification of ceramide on antigenicity of the carbohydrate of ganglioside, the reactivity of O-acetyl GM3 having 3-O-acetyl ceramide, which has been characterized as a glioma-related ganglioside, with monoclonal antibody M2590 was examined in comparison to that of non-acetylated GM3, by means of quantitative enzyme-linked immunosorbent assay, TLC-immunostaining and liposome immune lysis assay. In all these assay systems, O-acetyl GM3 showed less activity than GM3 as follows: GM3 was detected till 0.1 nmol in TLC-immunostaining, whereas O-acetyl GM3 could not be detected even at 0.25 nmol; the GM3 reaction was approximately twofold that of O-acetyl GM3 at each diluted point in the enzyme-linked immunosorbent assay; and 20% of the liposomes containing GM3 were lysed at 6 mol%, while liposomes containing O-acetyl GM3 did not lyse at that concentration. The lesser antigenicity of the sugar moiety of O-acetyl GM3 could be ascribed to the presence of an acetyl group in the ceramide at the 3-position of sphingosine.

The abundant and restricted expression of surfactant proteins SP-A and SP-D within the lung makes these collectins specific markers for lung diseases. The measurement of SP-A and SP-D in amniotic fluids and tracheal aspirates reflects lung maturity and the production level of the lung surfactant in infants with respiratory distress syndrome (RDS). The SP-A concentrations in bronchoalveolar lavage (BAL) fluids are significantly decreased in patients with acute respiratory distress syndrome (ARDS) and also in patients at risk to develop ARDS. The prominent increase of these proteins in BAL fluids and sputum is diagnostic for pulmonary alveolar proteinosis (PAP). The concentrations of SP-A and SP-D in BAL fluids from patients with idiopathic pulmonary fibrosis (IPF) and interstitial pneumonia with collagen vascular diseases (IPCD) are rather lower than those in healthy controls and the SP-A/phospholipid ratio may be a useful marker of survival prediction. SP-A and SP-D appear in the circulation in specific lung diseases. Their serum concentrations significantly increase in patients with PAP, IPF and IPCD. The successive monitoring of serum levels of SP-A and SP-D may predict the disease activity. The serum SP-A levels increase in patients with ARDS. SP-A is also a marker for lung adenocarcinomas and can be used to differentiate lung adenocarcinomas from other types and metastatic cancers from other origins, and to detect metastasis of lung adenocarcinomas.

To determine O-esterified positions, a rapid and complete acetalization to prepare an intermediate was established using ethyl vinyl ether as a new reagent. The new method was applied to O-esterified glycolipids followed by GC-MS analysis of the monosaccharide derivatives after methylation and methanolysis, revealing the derivatives with correctly substituted positions. This method was superior in terms of its shorter reaction time and complete acetalization, particularly of the N-glycolyl hydroxyl residue, to previously reported methods using methyl vinyl ether.

A sulfatide, O-fatty-acylated 3-sulfogalactosylceramide at C6-O on galactoside, was isolated from equine brain and the chemical structure was characterized by proton NMR and MS. The O-acylation site of the acylated sulfatide was determined by the down-field shift of protons attached to a carbon having an O-acyl group in the NMR spectrum and by analysis of a partially methylated derivative before and after acetalization of the intact sulfatide using GC-MS. The O-acyl chain length was determined by GLC, revealing that it exclusively had palmitoyl and stearoyl residues as the major fatty acids. The enzymatic conversion to the O-acyl sulfatide was further examined using equine brain microsomes as an enzyme source and different lipid substrates, resulting in O-acylation of 3-sulfogalactosylceramide from stearoyl CoA, while 6-O-acyl galactosylceramide was not O-sulfated from phosphoadenosine phosphosulfate. The results were supported by the comparably different N-linked fatty acid components between two lipid substrates, in which the component of 6-O-acyl sulfatide was mostly similar to that of sulfatide, but not to 6-O-acyl galactosylceramide.

To study the healing process of vascularized intercalary bone allograft after withdrawal of immunosuppressive drugs, allotransplantation of the tibia diaphysis with a vascular pedicle was performed in eight adult mongrel dogs (group 2) and assessments were made both during administration and after discontinuation of cyclosporin A. As controls, similar grafts with the vascular pedicles were removed and reimplanted back to the same animals (five dogs, group 1). Allotransplantation of frozen stored bone without a vascular pedicle (10 dogs, groups 3A and 3B) were also compared. No union occurred in most cases of frozen stored bone allotransplant because the transplanted bone was resorbed, leading to loosening and subsequent failure of osteosynthesis with the plate and screws used. Under cyclosporin A immunosuppression, bony union (i.e., when trabeculae were seen crossing the graft-recipient junction with obliteration of the junction line) occurred at almost similar time intervals in all dogs of group 2 (bone allotransplant with a vascular pedicle) by 3 months postoperatively, which was similar to those of group 1. No systemic side effects of cyclosporin A were observed. Cyclosporin A was discontinued 3 months following graft implantation. The bone graft became avascular within a week following withdrawal of cyclosporin A. However, bone union was maintained, and the transplanted bone never showed bone resorption, sclerosis, or fracture on serial radiographs up to the time the animals were sacrificed, between 5 and 14 months later. Histology at sacrifice showed that the transplanted allografts were being replaced at both ends by fresh bone derived from the transplantation bed. We conclude on the basis of the results of this study that solid bony union can be obtained in allotransplanted bone with a vascular pedicle if cyclosporin A is given for a brief period. After cyclosporin A is withdrawn, although the bone becomes nonviable secondary to rejection occurring in the blood vessels, its skeletal structure remains intact, enabling it to maintain its structural support while awaiting replacement by bony ingrowth from both ends of the graft.

Five microbial strains that removed hydrogen sulfide (H2S) or methylmercaptan (CH3SH) gas were newly isolated from soil samples. Strain DO-1, one of the isolates, was identified as a member of Pseudomonas sp., and it's immobilized cells removed 1 or 10 ppm of H2S gas within 2 hours. When strain DO-1 was cultured aerobically in a flask containing nutrient broth medium, the deodorizing activity increased, depending on the growth of the culture, and the maximum activity was obtained after 48 hours. Even though the immobilized cells were stored at 4 or 25°C in sealed bottles for 6 months, the deodorizing activity remained. Throughout this study, strain DO-1 removed H2 S gas without preliminary feeding or exposure to sulfur com-pounds as growth substrates or inducers. These characteristics are advantageous for the deodorization of the malodorous gases surrounding us in daily life.

We studied the individual and synergistic effect of 3 immunosuppressive drugs, FK-506 (1 mg/kg/day), 15-deoxyspergualin (2.5 mg/kg/day) and cyclosporine (15 mg/kg/day) in a DA/Lewis rat limb allotransplantation model. 74 right hindlimb transplantations were performed. The median time for onset of rejection was 4 days in animals without immunosuppression, 37 days in animals receiving cyclosporine immunosuppression for 30 days, 61 days in animals receiving FK-506 for 30 days, 36 days in animals receiving a 30-day course of cyclosporine and, in the first 15 days, a course of 15-deoxyspergualin, and 76 days in animals receiving a 30-day course of FK-506 and 15-deoxyspergualin in the first 15 days. The combination of cyclosporine with 15-deoxyspergualin did not prolong graft survival and no synergistic effect was evident. In contrast, survival time in rat limb allografts receiving FK-506 and 15-deoxyspergualin was longer than in those receiving single FK-506 therapy. Our findings suggest a positive synergistic immunosuppressive effect with FK-506 and 15-deoxyspergualin in limb allotransplantation.

Mannose-binding protein (MBP) belongs to the collectin subgroup of C-type lectins with specificity for mannose and N-acetylglucosamine sugars. We investigated whether rat MBPs isolated from serum (S-MBP) and liver (L-MBP) interact with phospholipids using antibody against each MBP. Both S- and L-MBPs bound to phosphatidylinositol coated onto microtiter wells in a concentration- and a Ca2+-dependent manner. L-MBP also bound to phosphatidylglycerol and weakly to phosphatidylserine. MBPs interacted with liposomes composed of these lipids. S- and L-MBPs bound to phosphatidylinositol 4-monophosphate. L-MBP also bound to cardiolipin. These results provide evidence for a novel type of ligand binding specificity for MBPs, and raise the possibility that phospholipids are ligands for collectins.