Measurements of Strong-Interaction Effects in Kaonic-Helium Isotopes at Sub-eV Precision with X-Ray Microcalorimeters.

We have measured the 3d→2p transition x rays of kaonic ^{3}He and ^{4}He atoms using superconducting transition-edge-sensor microcalorimeters with an energy resolution better than 6 eV (FWHM). We determined the energies to be 6224.5±0.4(stat)±0.2(syst) eV and 6463.7±0.3(stat)±0.1(syst) eV, and widths to be 2.5±1.0(stat)±0.4(syst) eV and 1.0±0.6(stat)±0.3(stat) eV, for kaonic ^{3}He and ^{4}He, respectively. These values are nearly 10 times more precise than in previous measurements. Our results exclude the large strong-interaction shifts and widths that are suggested by a coupled-channel approach and agree with calculations based on optical-potential models.

Feasibility and Outcomes of Direct Dual Portal Vein Anastomosis in Living Donor Liver Transplantation Using the Right Liver Graft With Anatomic Portal Vein Variations.

BACKGROUND: Portal vein (PV) reconstruction is a crucial factor in successful living donor liver transplantation (LDLT). In LDLT using the right liver grafts with anatomic PV variations, we sometimes encounter dual PV anastomosis. In this study we describe PV variations of donor liver in detail as well as our experiences with PV reconstruction in right liver grafts with PV variations. METHODS: We performed LDLT in 149 recipients between 2002 and 2016. PV variations of donor liver were classified into 3 major anatomic patterns, and we retrospectively analyzed the procedure and postoperative complications of PV anastomosis. RESULTS: PV variations in donor livers were classified as type A (normal type) in 125 patients, type B (trifurcation type) in 7 (4.7%), and type C (caudal origin of the right posterior branch) in 17 (11.4%). Among 75 right liver grafts, 10 (13.3%) had anatomic PV variations. In 9 of 10 recipients, dual PV of the graft were anastomosed to dual PV branches of the recipient in direct end-to-end fashion. In the remaining recipient, the posterior portal branch of the graft was anastomosed to the recipient portal trunk through the interposed venous graft in end-to-end fashion and the anterior portal branch of the graft was anastomosed to the side wall of the interposed venous graft. These 10 recipients did not develop any postoperative complications associated with PV anastomosis, although 3 of the 149 recipients (2.0%) developed complications associated with PV anastomosis, such as thrombosis and necrosis. CONCLUSION: Dual PV anastomosis of the right liver graft is safe and feasible in LDLT, even in anatomic PV variations.

Early Enteral Feeding of Daikenchuto Stimulates Early Bowel Movement With Increased Portal Venous Blood Flow After Living Donor Liver Transplantation.

Daikenchuto (DKT), a Japanese Kampo medicine, had been reported to increase small intestinal blood flow after liver resection. The aim of this study was to evaluate the effects of early enteral feeding of DKT on portal venous flow and early bowel movement after living donor liver transplantation (LDLT) in an attempt to clarify whether these effects on bowel motility can prevent bacterial and/or fungal translocation. MATERIALS AND METHODS: Our prospective study included the consecutive 16 LDLT recipients at Mie University Hospital between June 2006 and September 2009. Sixteen patients were divided into the 2 groups according to enteral feeding starting postoperative day (POD) 1: 8 patients in DKT (15 g/d) administration (DKT group, for 1 week) and 8 patients in tepid water administration (non-DKT group, for 1 week). Portal venous flow, portal venous pressure, presence of fungal infection (serum level of ß-D-glucan and fungal polymerase chain reaction assay), time to first food intake, and time to first defecation were serially examined. RESULTS: Portal venous flow (mean [SD] velocity) was significantly increased in DKT group compared with non-DKT group: 47.5 (12.9) vs 31.8 (15.4) (P = .04) on POD 1, 46.8 (11.5) vs 28.8 (12.5) (P = .03) on POD 3, and 42.3 (17.2) vs 25.2 (9.0) (P = .05) on POD 5. However, mean (SD) portal venous pressures did not significantly change between the 2 groups. Between the 2 groups (DKT vs non-DKT), the day of first oral intake was not significantly different: 6.9 (2.5) vs 11.3 (8.7) (P = .061), but the mean (SD) day of first defecation was significantly shorter in the DKT group: 3.9 (1.1) vs 5.5 (2.6) (P = .02). Although fungal polymerase chain reaction assay was not significantly different between the 2 groups (4 vs 4 positive cases), the mean (SD) serum levels of ß-D-glucan were significantly lower in the DKT group than in the non-DKT group: 9.0 (7.4) vs 18.4 (15.9) pg/mL (P = .04). CONCLUSION: Early enteral feeding of DKT after LDLT increased portal vein blood flow without increasing portal vein pressure and stimulated early bowel movement, which in turn might prevent fungal translocation.

De Novo Malignancy Following Adult-to-Adult Living Donor Liver Transplantation Focusing on Posttransplantation Lymphoproliferative Disorder.

OBJECTIVE: In patients with living donor liver transplantation (LDLT), late-onset complications sometimes develop because of long-term use of immunosuppressive drugs. One of the immunosuppressive drug-related complications is de novo malignancies resulting in reduced survival. PATIENTS AND METHODS: Among 153 patients undergoing LDLT, we retrospectively reviewed the medical records of 97 adult recipients (February 2002 to May 2017), who had been followed-up at our hospital for more than one year after LDLT. The median age was 52 years old (20-70) and the median observational period was 6.9 years (2.4-15.3). RESULTS: De novo malignancy after adult LDLT developed in 11.3% (11/97) of patients, including posttransplantation lymphoproliferative disorder (PTLD) (n = 4) (2 in the brain and 2 in abdominal lymph nodes), lung cancer (n = 1), pancreatic cancer (n = 1), gastric cancer (n = 1), laryngeal cancer (n = 1), lower gingival cancer (n = 1), bladder cancer (n = 1), and melanoma (n = 1). Age at cancer diagnosis ranged from 36 to 70 years old with an average age of 61 years. The interval from LDLT to cancer diagnosis was 8.3 years (3.9-12.2). Four patients (36.6%) including PTLD (n = 2), lung cancer (n = 1), and pancreatic cancer (n = 1) died of cancer and all of them were diagnosed with cancer within 10 years after LDLT. Six patients were diagnosed with cancer more than 10 years after LDLT and all of them survived after treatment of cancer. CONCLUSION: De novo malignancy was found in 11.3% of LDLT patients, and more than half of this population subset developed tumors 10 years after LDLT. Long-term close follow-up should be performed by taking any kinds of de novo malignancy into consideration.

Biliary Complications During and After Donor Hepatectomy in Living Donor Liver Transplantation Focusing on Characteristics of Biliary Leakage and Treatment for Intraoperative Bile Duct Injury.

BACKGROUND: Biliary complication is one of the major donor complications during and after hepatectomy in living donor liver transplantation (LDLT). We evaluated risk factors for donor biliary complication in adult-to-adult LDLT. PATIENTS AND METHODS: From March 2002 to November 2016, 126 consecutive patients who underwent donor hepatectomy in adult-to-adult LDLT were divided into 2 groups according to biliary compilations: nonbiliary complication (non-BC) group (n = 114) and biliary complication (BC) group (n = 12). RESULTS: Among 126 donor hepatectomies, 35 patients (28%) experienced perioperative complications, including 10 (7.9%) with Clavien-Dindo classification grade III. Biliary complications occurred in 12 patients (9.5%): bile leakage in 10 and intraoperative bile duct injury in 2. Additional computed tomography- and/or ultrasound-guided drainage or exchange of original drain was required in 7 patients. In comparison between BC and non-BC groups, future remnant liver volume was significantly higher in the BC group than in the non-BC group (63% vs 40%; P = .02). In multivariate analysis, larger future remnant liver volume (P = .005) and shorter operating time (P = .02) were identified as independent risk factors for biliary complications. We had 2 patients with intraoperative bile duct injury: both were successfully treated by duct-to-duct biliary anastomosis with insertion of biliary stent or T-tube. CONCLUSION: Large remnant liver volume was a significant risk factor for biliary complications, especially biliary leakage, after donor hepatectomy. For intraoperative bile duct injury, duct-to-duct anastomosis with biliary stent is a feasible method to recover.

Recombinant Human Soluble Thrombomodulin Attenuates Hepatic Ischemia and/or Reperfusion Injury by Inhibiting Leukocyte Accumulation in Mice With Normal and Fatty Liver.

In an attempt to increase the number of donor livers, there has been an increased use of marginal donor livers, such as steatotic (fatty) livers that increase susceptibility to ischemia and reperfusion injury (IRI). Inflammatory cell accumulation has a greater role in IRI in steatotic liver than in normal liver. Although the recombinant human soluble thrombomodulin (rhsTM) attracts attention as a new treatment for disseminated intravascular coagulation, the therapeutic efficacy of rhsTM in hepatic IRI remains uncertain, especially in fatty livers. We aimed to demonstrate the effect of rhsTM on hepatic IRI using well-established in vivo experimental models with steatotic liver. METHODS: C57/BL6 mice were divided into 2 groups: normal liver (NL) group and fatty liver (FL) group, in which the steatotic liver was induced by high-fat diet for 9 weeks. The mice in the NL and FL groups were premedicated with venous injection of rhsTM (TM) or saline (Control) as control groups. All 4 groups (NL-Control vs NL-TM, FL-Control vs FL-TM) were subjected to partial hepatic warm ischemia followed by reperfusion. RESULTS: rhsTM significantly attenuated liver injury in the FL group as well as the NL group, as evidenced by transaminase levels and histologic finding after hepatic IRI. rhsTM remarkably decreased the accumulation of inflammatory cells, such as macrophages and neutrophils, in both NL and FL tissue after IRI. Furthermore, rhsTM depressed mRNA and protein expressions of adhesion molecules such as intracellular adhesion molecule-1 and vascular cell adhesion molecule-1 in both NL and FL groups after IRI. CONCLUSION: Our results demonstrate that rhsTM has a protective effect on fatty liver as well as normal liver after hepatic IRI. They also suggest that rhsTM contributes to attenuation of leukocyte accumulation caused by depressing expressions of adhesion molecules that facilitate accumulation of leukocytes in liver tissue in hepatic IRI.

Isolated Biliary Fistula After Donor Right Hepatectomy and Its Novel Interventional Treatment: Isolated Liver-Punctured Drainage.

Isolated biliary leakage is difficult to manage, and afflicted patients often develop refractory fistula. The present case was a 43-year-old male donor whose wife developed acute fulminant liver failure. Computed tomography (CT) volumetry showed that the estimated remnant liver volume was only 394 mL (31%) if his right lobe would be harvested. Since remnant liver volume was marginal, our proposed cut line for the right hepatectomy was set in order to preserve branches of the middle hepatic vein draining segments 4b+8 and 5. Right hepatectomy was performed, but on postoperative day 14, the donor developed fever and right back pain, and enhanced CT showed a 6 cm intra-abdominal abscess at the site of cutting, and we diagnosed it as an isolated biliary fistula since the isolated segment 5/8 was receiving arterial blood supply and exhibiting regrowth. A transabdominal abscess drainage was performed, after which the patient lost 30 to 50 mL of bile juice per day in drainage until 2 months after the drainage procedure. Ethanol injection, acetic acid injection, and transarterial or portal embolization for the isolated liver were proposed, but these all were impossible to carry out because there were no accessible routes. Thus, re-abscess drainage with a 7-French drainage catheter was performed through the isolated liver on postoperative day 53, and the isolated functional liver was punctured to induce liver atrophy. After this drainage, the isolated liver started to shrink and bile output had been stopped. In conclusion, our punctured-liver drainage could be effective for the treatment of isolated biliary fistula, allowing physicians to avoid an invasive additional liver resection or other invasive percutaneous approach using chemical reagents.

Intravenous Administration of Tacrolimus Stabilizes Control of Blood Concentration Regardless of CYP3A5 Polymorphism in Living Donor Liver Transplantation: Comparison of Intravenous Infusion and Oral Administration in Early Postoperative Period.

BACKGROUND: We compared achievement rate of sufficient tacrolimus blood concentration in the early postoperative period and incidence of acute cellular rejection within 1 month after living donor liver transplantation (LDLT) between tacrolimus intravenous (IV) and oral administration groups. METHODS: From October 2005 to November 2016, 61 LDLT patients administered tacrolimus, who could be genotyped for CYP3A5*3 and *1, were chosen from the electronic record database. The patients were then divided into the 2 groups (an IV group [n = 38] and an oral group [n = 23]). We defined patients with 1*1 or *1*3 as expressors and those with *3*3 as nonexpressors. Sufficient trough level tacrolimus blood concentration on postoperative day (POD) 3 was defined as 10-20 ng/mL. RESULTS: Comparable concentrations were seen between the 2 groups, with mean blood concentration 13.7 ± 8.5 ng/mL in the oral group and 15.2 ± 4.3 ng/mL in the IV group. Achievement rate of sufficient tacrolimus concentration on POD 3 was significantly higher in the IV group than in oral group: 97% (37 of 38) vs 65% (15 of 23), respectively (P = .001). When we focused on achievement rate in the oral group according to CYP3A5 polymorphism, the frequency of expressors (17%) was significantly lower than that of nonexpressors (82%) (P = .016). However, in the IV group this negative influence was totally eliminated, resulting in high achievement rates regardless of CYP3A5 polymorphism. In terms of incidence of acute cellular rejection, there was no significant difference between the 2 groups (IV 32% vs oral 17%, P = .250). CONCLUSION: IV administration of tacrolimus allowed us to obtain more stable control of blood concentration regardless of CYP3A5 genotype.

Pretransplant Serum Hyaluronic Acid Can Be a Biomarker as a Prognostic Predictor in Adult-to-Adult Living Donor Liver Transplantation.

OBJECTIVE: The goal of this study was to evaluate whether pretransplant serum hyaluronic acid (HA) levels can predict outcomes after adult-to-adult living donor liver transplantation (LDLT). METHODS: In study I, 21 patients who underwent LDLT (March 2002-February 2004) were divided into 2 groups: the H-I group (HA ≥500 ng/mL; n = 12) and the L-I group (HA <500 ng/mL; n = 9). The influence of pretransplantation HA levels on short-term surgical outcome was investigated. In study II, 77 LDLT patients (May 2004-December 2014) were also divided into 2 groups: the H-II group (HA ≥500 ng/mL; n = 40) and the L-II group (HA <500 ng/mL; n = 37). We compared long-term survival and investigated prognostic factors. RESULTS: In study I, HA levels significantly decreased after LDLT, and those in the H-I group were significantly higher compared with the L-I group at 1, 3, 5, and 7 days after LDLT. There were significant differences in postoperative peak total bilirubin levels (H-I vs L-I, 17.2 vs 6.2 mg/dL; P = .013), peak ascitic fluid volume (1327 vs 697 mL/d; P = .005), and the hepatocyte growth factor levels at 3 days after LDLT (1879 vs 1092 pg/mL; P = .03). In study II, the 1- and 5-year survival rates were significantly lower in the H-II group than in the L-II group (H-II vs L-II, 65.0% and 48.5% vs 86.5% and 80.8%; P = .004). In multivariate analysis, significant prognostic factors were preoperative HA ≥500 ng/mL (P = .004) and graft to recipient body weight ratio <0.8 (P = .042). CONCLUSIONS: Preoperative HA level can be a prognostic risk factor. Patients with high HA levels are vulnerable and should be carefully managed after LDLT.

Living Donor Liver Transplantation for Biliary Atresia With Severe Preduodenal Portal Vein Stricture: Success and Pitfall of Portal Vein Reconstruction.

BACKGROUND: We report a rare case of 10-month-old female who underwent living donor liver transplantation (LDLT) for syndromic biliary atresia with preduodenal portal vein (PV) and its severe stricture owing to the previous Kasai portoenterostomy. Because we successfully performed "left at right liver transplantation (LAR-LT) and graft rerotation" in this case, we are present tips and pitfalls for this operation. METHODS: Preoperative computed tomography scan showed that her preduodenal PV was stenotic from the confluence of the superior mesenteric vein and splenic vein to hepatic hilum, which made us consider the necessity of ≥3 cm interposition vein graft to complete a safe PV anastomosis. To reduce a gap between donor and recipient's PV, we decided to put a left lateral section graft at the right subphrenic space called left-at-right liver transplantation. Thus, LDLT was performed with an identical lateral sectional graft from her father. After total hepatectomy, we implanted a graft in her right subphrenic space, and anastomosed the donor left hepatic vein to her inferior vena cava. Then, we anastomosed an interposition graft harvested from her left internal carotid vein to her PV. RESULTS: Even after reflowing PV flow, because the duodenum compressed the interposition vein graft, PV flows were totally insufficient. Therefore, we flipped a liver graft 180° from right to left upper abdominal cavity, which could reduce the gap between PVs and acceptable PV flow was obtained. CONCLUSIONS: In the present case, LAR-LT could reduce the distance of PVs. In addition, our rerotation method could be useful to alleviate tension on the PV anastomosis caused by preduodenal PV.

Impact of Splenectomy Just Before Partial Orthotopic Liver Transplantation Using Small-for-Size Graft in Rats.

BACKGROUND: Portal hypertension is a serious obstacle of partial orthotopic liver transplantation (POLT) with the use of small-for-size liver graft. Several therapeutic strategies including surgical innovations and pharmacological agents to reduce the portal hypertension have been developed. Splenectomy (SP) on POLT is one of surgical procedures to reduce portal pressure. We previously reported a dual cytoprotective mechanism of SP just before POLT, using small-for-size liver graft in a rat model. However, the best timing of SP during POLT has been unclear. We compared liver functions between SP just before and after POLT, using small-for-size rat liver grafts. METHODS: With the use of small-for-size liver grafts (20%) in rats previously reported, the rats were assigned to 2 groups: the pre-SP group (SP just before POLT) and the post-SP group (SP just after POLT). Liver tissues and blood were sampled at 6 and 24 hours after POLT for several liver function tests. RESULTS: The serum alanine aminotransferase levels at 24 hours after POLT were significantly decreased in the pre-SP group compared with the post-SP group (226 ± 78 vs 340 ± 71 IU/L). The infiltrations of neutrophil at 6 hours and ED-1-positive cells at 24 hours were significantly suppressed in the pre-SP group compared with the post-SP group. Serum hyaluronic acid levels, indicating attenuation of endothelial damage, were lower in the pre-SP group than in the post-SP group. CONCLUSIONS: SP before POLT, which directly eliminates splenic inflammatory leukocytes, inhibits inflammatory leukocyte infiltration, which leads to impaired liver function as compared with SP after POLT.

Influence of Angiotensin-converting Enzyme Genetic Polymorphism on Late Renal Dysfunction After Adult-to-adult Living-donor Liver Transplantation.

BACKGROUND: Late renal dysfunction (LRD) is known to be one of the most important complications to affect long-term outcome after living-donor liver transplantation (LDLT). The relationship between angiotensin-converting enzyme insertion (I)/deletion (D) gene polymorphism and renal function after LDLT are still unknown. The aim of this study was to elucidate the risk factors for LRD after LDLT, focusing on ACE gene polymorphism. MATERIALS AND METHODS: Among the 94 recipients who underwent adult-to-adult LDLT between March 2002 and September 2009, the total number of subjects who survived more than 1 year after LDLT and in whom angiotensin-converting enzyme genotype could be measured was 64. LRD was defined as estimated glomerular filtration rate level less than 60 mL/min/1.73 m(2) at any point after 1 year from undergoing LDLT. RESULTS: LRD was found in 24 patients (37.5%). The incidence of LRD was significantly higher in D/D type than in I/I or I/D type: 85.7% (6/7) vs. 42.1% (8/19), 35.7% (10/38) (P = .010). Preoperative estimated glomerular filtration rate was significantly lower in D/D type than in I/I, I/D types, and postoperatively they were significantly lower in D/D type at 2, 3, and 4 years after LDLT. By multivariate analysis, age and hypertension were the independent risk factors for LRD. The 10-year survival rate was much lower in the recipients with LRD than in those without LRD at 66.7% versus 87.5%, respectively (P = .053). CONCLUSION: In conclusion, age and hypertension were determined as significant independent risk factors for LRD after adult-to-adult LDLT, and the recipients with D/D genotype should be strictly cared for the development of LRD.

Effects of administration of murine recombinant IL-4 on the resistance of mice to Listeria monocytogenes infection.

Mice given recombinant murine interleukin-4 as a single i.v. bolus concomitant with a Listeria monocytogenes challenge did not display increased anti-listeria resistance. Under certain conditions, IL-4 administration slightly increased the bacterial burdens in the spleens and livers of infected mice. This finding is consistent with previous reports that endogenous IL-4, or transfer of IL-4 producing TH2 cells, can be detrimental to host defense against microbial infection.

Establishment of a CCR5-expressing T-lymphoblastoid cell line highly susceptible to R5 HIV type 1.

The beta-chemokine receptor CCR5 is considered to be an attractive target for inhibition of CCR5-using (R5 or macrophage-tropic) HIV-1. However, R5 HIV-1 cannot replicate in CD4+ T cell or monocyte lines because of the lack of CCR5 expression on their surface, which apparently hampers discovery and development of effective CCR5 antagonists against HIV-1 replication. In this study, we have established the CCR5-expressing T cell line MOLT-4/CCR5, highly permissive to the replication of R5 HIV-1. The cells express a considerable amount of CCR5 on their surface. When the cells were infected with the R5 HIV-1 strains Ba-L and JR-FL, the virus-induced cytopathic effect (syncytium formation) was observed, and the cells produced large amounts of HIV-1 p24 antigen in the culture supernatants. The analyses of progeny viruses for their coreceptor use and gp120 V3 nucleotide sequence revealed that they were R5 HIV-1. The parental cell line MOLT-4 was much less susceptible to Ba-L and totally insusceptible to JR-FL. Furthermore, MOLT-4/CCR5 cells could support the replication of an R5 clinical isolate, but MOLT-4 cells could not. When TAK-779, a novel small-molecule nonpeptide CCR5 antagonist, was examined for its inhibitory effect on R5 HIV-1 replication in MOLT-4/CCR5 cells, the compound displayed potent antiviral activity, as demonstrated in peripheral blood mononuclear cells. These results indicate that the established cell line will be an extremely useful tool for experiments with R5 HIV-1.

Streptococcus pneumoniae in the nasal cavity of mice causes lower respiratory tract infection after airway obstruction.

A model of persistent colonisation in the nasal cavity of mice by Streptococcus pneumoniae has been established. S. pneumoniae NNP-4 was introduced to the lungs of CBA/J mice at a density of c. 10(4) cfu/lung by an aerosol method and a high dose of ampicillin was administered 1 h after infection. This antibiotic eliminated bacteria from the lungs and trachea, but did not affect the bacterial counts in the nasal cavity. In mice given ampicillin, the bacteria were recovered from the nasal cavity only more than 2 weeks after infection, but IgG antibody against the colonising organisms was produced in sera around day 8 after infection. Airway obstruction was induced by intratracheal injection of formalin 2% into mice. Organisms appeared in the lungs in greater numbers when formalin was injected before the antibody production than when the immunity was established. In the early stages of infection, 10(3)-10(4) cfu appeared in the lungs 6 h after the formalin injection and the bacterial counts increased to c. 10(6) cfu within 24 h. When ampicillin was administered again 1 h after formalin was given, no bacteria were recovered from lungs 6 h later. However, in some of the mice given ampicillin after formalin, bacteria appeared in the lungs on the next day and the bacterial counts increased thereafter. These results suggest that S. pneumoniae in the nasal cavity invade the lower respiratory tract and that these organisms can localise and proliferate in lungs in the event of damage to the airway.

Establishment of a model of penicillin-resistant Streptococcus pneumoniae pneumonia in healthy CBA/J mice.

Examination of strain differences in the susceptibility of mice to experimental respiratory tract infection with penicillin-resistant Streptococcus pneumoniae TUM19 revealed that a fatal infection model could be induced in immunocompetent CBA/J mice, but not in C3H/HeN, C57BL/6 or ICR mice. After intranasal instillation of c. 10(6) cfu of S. pneumoniae, the bacterial counts in the lungs of CBA/J mice increased from 10(5) to 10(7) cfu after 3-5 days, and gradually increased thereafter. The challenge organisms localised mainly in the lungs until 14 days after infection. Mice began to die c. 7 days after infection, and by 3 weeks most of the mice had died. Histopathologically, infiltration of neutrophils and lymphocytes around bronchi was observed from 1 day after infection, and fibrin deposition was seen in alveolar and bronchial spaces from 5 days. This model may be useful for investigating therapy of respiratory tract infection caused by penicillin-resistant S. pneumoniae because its pathological features resemble those observed in the human disease.

Chromosome analysis of human spermatozoa exposed to antineoplastic agents in vitro.

We studied in vitro the cytogenetic effects of six antineoplastic agents, bleomycin (BM), cyclophosphamide (CP), daunomycin (DM), methyl methanesulfonate (MMS), mitomycin C (MMC) and triethylenemelamine (TEM) on spermatozoa, using an interspecific in vitro fertilization system between zona-free hamster oocytes and human or bull spermatozoa. In preliminary experiments with bull spermatozoa, clastogenic effects were clearly shown with BM, DM, MMS and TEM, but not with CP and MMC. In main experiments, the effects of the first four chemicals were studied in detail with human spermatozoa. Total numbers of 585 and 512 spermatozoa were karyotyped in the control and the chemical-treated groups respectively. The incidence of spermatozoa with structural chromosome aberrations was 34.5%, 53.0%, 59.3%, and 55.6% in the BM (50 micrograms/ml, 90 min), DM (0.1 microgram/ml, 90 min), MMS (100 micrograms/ml, 120 min) and TEM (0.1 micrograms/ml, 120 min) groups respectively, each showing a significantly higher incidence than the matched controls (10.1-13.5%). Breakage-type aberrations were more frequent than exchange-type aberrations in the BM, MMS and TEM groups, while the exchange-type aberrations were more frequent in the DM group. Exchanges were mainly of the chromatid type in the DM, MMS and TEM groups, while chromosome-type exchanges occurred more frequently in the BM group. These results are discussed in relation to previous data on chemical-induced chromosome aberrations in mammalian somatic cells and in mouse spermatozoa.

Studies on anti-MRSA parenteral cephalosporins. I. Synthesis and antibacterial activity of 7beta-[2-(5-amino-1,2,4-thiadiazol-3-yl)-2(Z)- hydroxyiminoacetamido]-3-(substituted imidaz.

In order to improve the antibacterial activity of cefozopran (CZOP) against methicillin-resistant Staphylococcus aureus (MRSA), we initiated chemical modification to introduce a 2-(5-amino-1,2,4-thiadiazol-3-yl)-2(Z)-hydroxyimino acetyl group at the C-7 position and a 3- or 6-substituted imidazo[1,2-b]pyridazinium or 5-substituted imidazo[1,2-a]pyridinium group at the C-3' position. Although this approach successfully enhanced the anti-MRSA activity of CZOP two to eight times, a slight decrease in the activity against Gram-negative bacteria including Pseudomonas aeruginosa was involved. Among the novel derivatives, 3-(6-aminoimidazo[1,2-b]pyridazinium-1-yl)methyl-7beta-[2-(5-amino-1,2,4-thiadiazol-3-yl)-2(Z)hydroxyiminoacetamido]-3-cephem-4-carboxylate (44a) showed an excellent balance of activity against MRSA and Gram-negative bacteria.

Studies on anti-MRSA parenteral cephalosporins. II. Synthesis and antibacterial activity of 7beta-[2-(5-amino-1,2,4- thiadiazol-3-yl)-2(Z)-alkoxyiminoacetamido]-3-(substituted imidaz.

In an effort to discover a novel cefozopran (CZOP) derivative having excellent antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA), we performed chemical modification of the alkoxyimino moiety and imidazo[1,2-b]pyridazinium group of CZOP. Among the prepared compounds, the cyclopentyloxyimino derivative 7beta-[2-(5-amino-1,2,4-thiadiazol-3-yl)-2(Z)-cyclopentyloxyiminoacetamido]-3-(3,6-diaminoimidazo[1,2-b]pyridazinium-1-yl)methyl-3-cephem-4-carboxylate (20 g) showed the most potent anti-MRSA activity, reflecting its high affinity (IC50 = 1.6 microg/ml) for penicillin binding protein 2' (PBP2'), although its anti-MRSA activity was slightly inferior to that of vancomycin (VCM). In experimental systemic infection in mice, however, 20 g showed activity comparable to that of VCM against MRSA. In addition, 20 g showed activity similar or slightly inferior to that of CZOP against Pseudomonas aeruginosa both in vitro and in vivo. Considering its favorable antibacterial activity profile, 20 g was considered to be the most promising CZOP derivative for further studies.

Studies on anti-MRSA parenteral cephalosporins. III. Synthesis and antibacterial activity of 7beta-[2-(5-amino-1,2,4-thiadiazol-3-yl)-2(Z)-alkoxyiminoacetamido-3.

In the course of our exploration for a novel cephalosporin derivative having excellent antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA), we modified the C-3 linked spacers of cephem derivatives bearing a 1-methylimidazo[1,2-b]pyridazinium-6-yl group at the C-3' position and 2-(5-amino-1,2,4-thiadiazol-3-yl)-2(Z)-cyclopentyloxy-iminoacetyl group at the C-7 position. The optimal spacers were the (E)-2-vinyl and (E)-2-thiovinyl groups seen in 19a and 29aa, respectively. Their anti-MRSA activity was 16 to 32 times as potent as that of cefozopran (CZOP). Focusing on the (E)-2-vinyl and (E)-2-thiovinyl spacers, we further modified the alkoxyimino groups in the C-7 acyl moiety and the 1-alkylimidazo[1,2-b]pyridazinium moieties at the C-3' position and investigated the structure-activity relationships (SAR) of the derivatives. Consequently, we selected 7beta-[2-(5-amino-1,2,4-thiadiazol-3-yl)-2(Z)-fluoromethoxyiminoacetamido]-3-[(E)-2-(1-methylimidazo[1,2-b]pyridazinium-6-yl)thiovinyl]-3-cephem-4-carboxylate (29ca) as a new anti-MRSA parenteral cephalosporin candidate for further biological evaluation. The selected 29ca showed anti-MRSA activity comparable to that of vancomycin (VCM) both in vitro and in vivo, high affinity (IC50)=2.7 microg/ml) for penicillin binding protein 2' (PBP2') of MRSA and potent activity against Gram-negative bacteria as well.