Small-angle synchrotron x-ray scattering reveals distinct shape changes of the myosin head during hydrolysis of ATP.

In the energy transduction of muscle contraction, it is important to know the nature and extent of conformational changes of the head portion of the myosin molecules. In the presence of magnesium adenosine triphosphate (MgATP), fairly large conformational changes of the myosin head [subfragment-1 (S1)] in solution were observed by small-angle x-ray scattering with the use of synchrotron radiation as an intense and stable x-ray source. The presence of MgATP reduced the radius of gyration of the molecule by about 3 angstrom units and the maximum chord length by about 10 angstroms, showing that the shape of S1 becomes more compact or round during hydrolysis of MgATP. Comparison with various nucleotide-bound S1 complexes that correspond to the known intermediate states during ATP hydrolysis indicates that the shape of S1 in a key intermediate state, S1-bound adenosine diphosphate (ADP) and phosphate [S1**.ADP.P(i)], differs significantly from the shape in the other intermediate states of the S1 adenosine triphosphatase cycle as well as that of nucleotide-free S1.

X-ray diffraction studies on the structural changes of rigor muscles induced by binding of phosphate analogs in the presence of MgADP.

To clarify the structure of the ATP hydrolysis intermediates (ADP.Pi bound state) formed by actomyosin crossbridges, the effects of various phosphate analogs in the presence of MgADP on the structures of the thin and thick filaments in glycerinated rabbit psoas muscle fibers in the rigor state have been investigated by X-ray diffraction with a short exposure time using synchrotron radiation. When MgADP and phosphate analogs such as metallofluorides (BeFx = 3,4 and AlF4) and vanadate (VO4(Vi)) were added to rigor fibers in the presence of the ATP-depletion backup system, the intensities of the actin-based layer lines were markedly weakened. The greatest effect (approximately 50% decrease in intensity) was observed in the presence of BeFx among the analogs examined. The intensity distribution of the 5.9 nm actin-based layer line shifted towards that observed in the Ca(2+)-activated fibers, while the first actin layer line at approximately 1/36.7 nm-1 retained a rigor-like profile with an intensity weakened by approximately 50%. The intensity of the equatorial 10 reflection increased while that of the 11 reflection changed little, resulting in only a small increase (approximately 1.7 fold) in the intensity ratio of the 10 to the 11 reflection. No resting-like pattern appeared upon the addition of MgADP and BeFx. These results indicate that a substantial fraction (approximately 40%) of the myosin heads dissociate from actin but the detached heads remain in the vicinity of the actin filaments when MgADP and BeFx bind. The states produced by binding phosphate analogs to a rigor muscle differ from the resting-like state produced by adding them to a contracting muscle (Takemori et al., J. Biochem. (Tokyo) 117 (1995) 603-608). Our conclusion put forward to explain the data is that one of the two heads of a crossbridge is detached and the other retains a rigor-like attachment.

Extensibility of the actin and myosin filaments in various states of skeletal muscle as studied by X-ray diffraction.

The effects of length changes applied to resting, contracting and rigor muscles on the reflection spacings of the X-ray diffraction patterns were summarized. The spacing changes of the actin- and myosin-based meridional reflections as a function of tension relative to an isometric tension of active muscle (P0) were linear and almost identical in the active and rigor states, showing that the extension of both filaments is Hookenian and does not depend upon the states of muscle. In addition to their length changes caused by tension generation, there are small but significant length changes of both filaments due purely to activation of muscle. The actin and myosin filaments are elongated by approximately 0.36% and approximately 0.43%, respectively under the maximum active tension. The results indicate that a large part of the sarcomere compliance of an active muscle is caused by the extensibility of the myofilaments. Inspection of the behavior of the meridional and layer-line reflection spacings reveals that there is a close relationship between the extensibility and helical twisting of the actin filaments under active and passive forces. The extension caused by tension is associated with an unwinding of right-handed helices following the actin monomers in the filament. At the pointed end of the filament could rotate anticlockwise through one fifth the complete turn during contraction.

Structure of the monomeric actin-myosin head complex as revealed by X-ray solution scattering.

In order to study the structure of a complex of monomeric actin and myosin head (S1) in the presence of nucleotide, non-polymerizable G-actin was prepared by the reaction of G-actin with m-maleimidobenzoic acid N-hydroxysuccinimide ester (MBS) and diazonium(1H)tetrazole (DHT). Although singly-modified MBS-actin forms a 2:1 complex with S1, doubly-modified DHT/MBS-G-actin forms a 1:1 complex with a dissociation constant of approximately 10(-6) M and can activate Mg-ATPase of S1 by 2-fold. Using a synchrotron X-ray source, we have measured X-ray solution scattering from reversible acto-S1 complexes obtained by mixing rabbit skeletal S1 and DHT/MBS-G-actin in the absence of nucleotide or in the presence of MgADP. The Guinier plots of the scattering intensity showed straight lines indicating no large aggregates and gave the radii of gyration of the complexes to be 49 and 46 A in the absence and presence of ADP, respectively. The analysis of the scattering curves showed that a monomeric actin binds to a tip of S1, and that the complex undergoes substantial rearrangement resulting in a more compact structure when it binds ADP.

Noncancerous pulmonary lesions in patients with metastatic testicular germ cell cancer.

By thoracoscopic surgery, we confirmed pulmonary noncancerous lesions in two patients with metastatic nonseminomatous testicular germ cell cancer. The purpose of thoracoscopic surgery was complete resection of postchemotherapeutic residual tumour in one patient or a biopsy for new lesions during chemotherapy. Histological findings were normal lymph node and pulmonary fibrosis. We demonstrated CT findings of these lesions and made some discussion on management of disseminated testicular cancer.

A case of adenocarcinoma of the renal pelvis.

A 28-year-old female had adenocarcinoma arising from the renal pelvis with ascites containing adenocarcinoma cells. The primary site was treated with radical nephrectomy, resection of the remnant ureter with a bladder cuff. Combination chemotherapy with cisplatin, doxorubicin, and cyclophosphamide (CAP) was performed as an adjuvant therapy. Approximately 3 years after the nephrectomy, she is currently alive with no clinical evidence of recurrence. CAP seems to have been effective in the treatment of the disease.

Lupus cystitis. report of a case.

A 42-year-old female with a history of systemic lupus erythematosus (SLE) visited the Department of Internal Medicine at our hospital complaining of anorexia. Hydronephrosis was diagnosed, and she was referred to our department for further evaluation. Intravenous pyelography (IVP) revealed bilateral hydronephrosis and hydroureters. Computerized tomography (CT) showed retention of ascitic fluid and thickening of the intestinal and bladder walls with contrast effects in the serosa and mucosa. Lasix-loaded renography showed that both kidneys were normal. Examination of a biopsy specimen revealed interstitial edema, fibrosis, and infiltration of inflammatory cells. Cystometry showed normal intravesical pressure. A diagnosis of lupus cystitis was made and administration of 40mg of prednisolone was started. The anemia was ameliorated, and thickening of the bladder and intestinal walls was no longer seen on CT. Hydronephrosis of the left kidney was less prominent. However, aggravation of the right ureteral stenosis was noted, and a stent was inserted in the stenosed ureter. Stenosis was successfully relieved by the stent, and internal drainage was established. The patient is still receiving steroids and is being followed at our department. Twenty-two reported cases of lupus cystitis in Japan are reviewed.

[Statistics of the operations at the Department of Urology, Tone Chuo Hospital during a six-year period (June 1993-May 1999)].

A clinical statistic survey was made on the operations performed at the Department of Urology, Tone Chuo Hospital between June 1993 and May 1999. The total number of operations was 1296, consisting of 97 (7.5%) operations of the kidney, 67 (5.2%) operations of the ureter, 190 (14.7%) operations of the bladder, 454 (35.0%) operations of the prostate, 63 (4.9%) operations of the urethra, 92 (7.1%) operations of the penis, 149 (11.5%) operations of the scrotum and 184 (14.2%) other operations. With the aging society, the number of operations for those over 60 years old has exceeded 60%.

Effect of intravesical instillation of antitumor drugs in the development of N-butyl-N-(4-hydroxybutyl) nitrosamine-induced bladder tumors in rats.

We examined the effects of intravesical instillation of adriamycin (ADM) and cis-diammine-dichloroplatinum (CDDP) on the development of bladder tumors induced by 0.05% N-butyl-N-(4-hydroxybutyl)nitrosamine in 115 rats. Intravesical instillation was performed in two ways; continuous administration (5 times a week for 5 weeks) and intermittent administration (once a week). Group 1 (control group) did not receive intravesical instillation. Group 2 I(ADM), group 3 (CDDP) and group 4 (physiological saline) were continuous type. Group 5 (ADM), group 6 (CDDP) and group 7 (physiological saline) received intermittent administration. Bladder weight was significantly higher in group 4 than in groups 1 or 7, and that in groups 2 and 3 than that in group 5 and 6. In groups 2, 3, 5 and 6 bladder weight was almost normal or higher than in the control group, and in group 3 histologically cancer was not seen in one rat. Physiological saline had promoting activity, and ADM and CDDP had both inhibitory and promoting activities. Also, intravesical instillation itself was suggested to promote tumor development under carcinogenic circumstances. We conclude that intermittent intravesical instillation should be performed to inhibit tumor recurrence and intravesical instillation therapy should not be performed clinically for a long period.

[Clinical analysis of ureteral injuries].

Six cases of ureteral injuries were analyzed. Two cases were male and 4 cases were female. The right side was affected in 1 case and the left in 5 cases. The affected portion was the uretero-pelvic junction in 1 case and lower iliac vessels in the other cases. Five cases were iatrogenic injuries in surgical or gynecological pelvic operation, and the other was due to blunt trauma. Intraoperative diagnosis was made in 4 cases. Drain leakage led to diagnosis in one case and pyelonephritis in another. Ureteroureterostomy was performed in 2 cases, pyeloureterostomy in 1 case, ureteroneocystostomy with psoas hitch in 1 case, Boari bladder flap in 1 case and simple suture in 1 case. 4-0 or 5-0 Dexon with an atraumatic needle was used for all anastomoses. Except for case 1, who died of the recurrence of carcinosarcoma, renal functions were preserved in all patients during the follow-up period ranging from 5 months to 7 years 8 months.

Pheochromocytoma of the urinary bladder: a case report.

We present a 66-year old female patient with pheochromocytoma of the urinary bladder. We performed transabdominal needle biopsy of the tumor without suspicion of pheochromocytoma because of her well-controlled blood pressure and no characteristic symptoms following administration of antihypertensive medication. Hypertensive crisis (260/130 mmHg) occurred just after the needle insertion. The diagnosis was pheochromocytoma. The norepinephrine level in the serum and her blood pressure normalized without antihypertensive medication after partial cystectomy. Pheochromocytoma should be suspected in cases of intramural bladder tumors, especially in a normotensive patients receiving antihypertensive medication.

[Clinical features of multiple primary cancers including prostate cancer].

We analyzed the clinical features of multiple primary cancers (MPCs) that included prostate cancer. MPCs were observed in 93 (15.2%) of the patients suffering from prostate cancer. In the MPC group, the organ most commonly involved was the stomach, followed by bladder, colon and lungs. The median age at diagnosis of the first, second and third cancers was 72, 74, and 75 years old, respectively, and the duration between the first and second cancers (median: 20 months) was longer than that between the second and third cancers (median: 8 months). In the 37 MPC patients whose cause of death was obvious, 29 (78.4%) died of a cancer; prostate cancer was not so common (6 patients) as the cause of death. Age at diagnosis and grade distribution of prostate cancer were not significantly different between the MPC and single primary cancer (SPC) groups. However, the proportion of earlier stage was significantly (p < 0.01) higher in the MPC group than in the SPC group, and this trend was more obvious in patients whose prostate cancer was diagnosed as the second cancer. The prostate cancer-specific survival rates were significantly higher in the patients with MPC, and this trend was more obvious in the patients with stage D or moderately differentiated cancer. It is important in the follow up of prostate cancer patients to be aware of the possibility of the occurrence of a second cancer.

[Clinical analysis of methicillin-resistant Staphylococcus aureus (MRSA) infection].

In 14 patients from which methicillin-resistant Staphylococcus aureus (MRSA) was isolated at the Department of Urology, Tone Chuo Hospital June, 1992 and October, 1993, the site of infection, background of patients, and drug resistance were analyzed. The 14 patients consisted of 11 males and 3 females between 45 and 85 years old with a mean of 71.6 years. The site from which MRSA was isolated was urine in 11, wound in 2, nasal cavity in 1, pharynx in 1, and renal fistula in 1 (detected at 2 or more sites in 2). The underlying condition was prostate hyperplasia in 2, prostate cancer in 4 (after radical prostatectomy in 1, complicated by bladder stone in 1), bladder tumor in 3, (during bladder instillation of BCG in 1), perirenal abscess in 2, renal pelvic tumor in 1, neurogenic bladder in 1, and after Boari's operation in 1. Urethral catheterization had been performed in 3. A fewer of 38 degrees C or above was noted in 3. Mixed infection was observed in 10, and was caused by Escherichia coli in 2, Proteus mirabiris in 1, Candida in 1, Klebsiella in 2, Pseudomonas aeruginosa in 2, and Serratia in 2. Four patients has previously been administered antibiotics, which were third generation cephems in 3 and penicillin in 1. The drug sensitivity was 100% for vancomycin (VCM), 30% for imipenam (IMP), 31% for minomycin (MINO), 31% for amikacin (AMK), and 7% for fosfomycin (FOM). As for chemotherapy, VCM+FOM+sulbactam/cefoperazone were administered to 6, and ceftazidime+ MINO were administered to 1.(ABSTRACT TRUNCATED AT 250 WORDS)

[Marked effectiveness of the LH-RH analogue in a case of prostate cancer with large lymph node metastasis].

A 78-year-old man visited our hospital complaining of pollakisuria, dysuria, and edema of lower extremities. Physical examination revealed a hard, fixed and fist size mass in the abdomen Lymph nodes of left supraclavicular fossa were hardly palpable. His prostate was larger than a hen's egg, stony hard and fixed in the pelvis on digital rectal examination (DRE). The prostate specific antigen (PSA) level was elevated to 584 ng/ml. Computerized tomography (CT) revealed enlarged retroperitoneal lymph nodes. Bone scan showed multiple abnormal uptake. Prostate biopsy showed poorly differentiated adenocarcinoma. Treatment with the LH-RH analogue was very effective. The retroperitoneal lymph nodes were no longer enlarged on CT. The prostate had become soft and was reduced to walnut size on DRE. The PSA level had decreased to within the normal range. The multiple abnormal uptake on the bone scan decreased. This is the 21st case of prostate cancer with large lymph node metastasis in Japan.

[Evaluation of transition zone biopsy in systematic prostate biopsy].

We evaluated the systematic biopsies performed on 83 patients suspected of having prostate cancer. In the systematic biopsy, 6 cores were from the peripheral zone and 2 cores from the transition zone. Cancer was detected in 25 patients (30.1%). The percentage of patients who had abnormal digital rectal examination and transrectal echo findings, average PSA and PSA density, and the number of examinations which suggested cancer were higher in the cancer group than in the non-cancer group, although the mean prostate volume was smaller. Cancer was more frequently detected in the peripheral zone than in the transition zone. Cancer was detected only in the transition zone in only 1 of the 25 cancer patients. We conclude that biopsy of the transition zone to all the patients is not always needed in systematic biopsy.

[Concentrations of new quinolone agents in serum and prostate tissue].

Twenty five patients with benign prostate hypertrophy were administered ofloxacin (OFLX) simultaneously with norfloxacin (NFLX) in 6 patients, ciprofloxacin (CPFX) in 10, tosfloxacin (TFLX) in 5 and enoxacin (ENX) in 4 patients. The dose of these new quinolones was 100 mg (except 150 mg of TFLX). Their concentrations in the serum and the prostate tissue were measured by high performance liquid chromatography (HPLC). The serum concentration of OFLX was significantly higher than that of NFLX, CPFX, TFLX or ENX. The prostate tissue concentration of OFLX was significantly higher than that of CPFX or TFLX. The ratio of tissue versus serum concentration of each new quinolone agent was not significantly different. The high OFLX tissue concentration appeared to be caused by the high serum concentration.

[Clinical evaluation of Augpenin (clavulanic acid-ticarcillin) in urinary tract infections: especially against beta-lactamase producing strains].

Augpenin contains ticarcillin (TIPC) and clavulanic acid (CVA), at a ratio of 15:1. The beta-lactamase inhibitor, CVA, has been added to protect the TIPC from inactivation by beta-lactamase. To investigate the drug efficacy and safety against urinary tract infections (UTI), Augpenin was administered to 33 patients with chronic complicated UTI and 7 patients with acute pyelonephritis. Thirty two cases were evaluable by the UTI criteria. Excellent results were obtained in 6 of the cases of acute pyelonephritis, and moderate results in 1 case, with an overall effectiveness rate of 100%. Excellent results were obtained in 14 of the cases of chronic complicated UTI, moderate results in 9 cases, and poor results in 2 cases, with an overall effectiveness rate of 92%. No adverse reactions were noted, but a transient elevation of glutamic oxaloacetic transaminase and glutamic pyruvic transaminase levels was observed in 2 cases.

[Clinical effect of Cernilton in chronic prostatitis].

Twenty-five patients with chronic prostatitis were given Cernilton tablets. Improvement of subjective symptoms and objective findings was noted in 96.0% and 76.0% of the cases. Sonographic findings in the prostate showed 33-100% improvement in four objective items. No side effects were observed in any case after Cernilton medication. Cernilton was judged to be an effective drug for chronic prostatitis.

[Measurement of serum PSA by DELFIA PSA kit and its application for mass screening. The Gunma Urological Oncology Study Group].

The significance of prostate specific antigen (PSA) measured by DELFIA PSA kit in the 1,177 serum samples examined by mass screening for prostate cancer was evaluated. All subjects were examined by questionnaire, digital rectal examination (DRE) and prostatic acid phosphatase (PAP) and the subjects in whom prostate cancer was suspected from abnormal DRE and/or elevated PAP were recommended to receive the secondary screening for the presence of prostate cancer. All serum specimens used for this study were kept in our serum bank. The cut-off value was investigated between non-cancer subjects (diagnosed as normal, voiding disturbance or BPH) and prostate cancer patients. When the cut-off value was 2.89 ng/ml, the sensitivity, specificity and accuracy as prostate marker was 80.6%, 90.0% and 89.4%, respectively. Therefore, the cut-off value was determined as 3.0 ng/ml. The significance of PSA was retrospectively evaluated compared to PAP in the subjects examined by our mass screening. Twenty eight of the 39 palpable prostate cancers which could not be detected from the PAP level were detected from the PSA level, namely the sensitivity of the detection using the PSA level is more excellent than that using PAP. However, the false negative rate obtained using PSA was high (30.3%) in the patients with stage B disease. The relationship among serum PSA concentration, prostate size estimated by DRE and age was investigated. PSA was increased with age and prostate size. This estimation should be confirmed by using an ultrasonography because the prostate size obtained by DRE is inaccurate as compared with that obtained by ultrasonography.

[PSA doubling time in prostate cancer relapsed after endocrine therapy].

PURPOSE: The PSA level of prostate cancer patients generally declines after endocrine therapy, but elevates when the cancer relapses in most cases. However, the rate of elevation differs with the case. We investigated the PSA doubling time (PSA-DT) of the prostate cancer patients whose PSA declined after endocrine therapy and later re-elevated, and investigated the relationship with other parameters. PATIENTS AND METHODS: We investigated 55 prostate cancer patients who underwent endocrine therapy between 1991 and 1998. Their PSA re-elevated continuously after their PSA fell below 10 ng/ml after the endocrine therapy as the first line treatment. First, the correlation coefficients with time and PSA were calculated in order to decide whether their PSA elevation was exponential or linear. PSA-DT was calculated thereafter, and compared with the clinical stage, pathological differentiation, clinical relapse style, time from the beginning of the therapy to PSA relapse, pre-treatment PSA value, and prognosis. The relationship between PSA-DT and each clinical parameter was tested using the Kruskal-Wallis test. Differences in survival rates and PSA-DT were calculated using the log-rank test. RESULTS: PSA elevated exponentially after cancer relapsed. PSA-DT in all cases ranged from 0.5 to 26.3 months, with an average of 4.4 +/- 4.8 (S.D.) months and the median was 2.5 months. PSA-DT was significantly (p < 0.01) short when the pre-treatment clinical stage was high, the time from the beginning of the therapy to PSA relapse was short, or the pre-treatment PSA value was high. PSA-DT tended to be short when the pre-treatment pathological differentiation was low, but not significantly. PSA-DT tended to be short when the cancer relapsed as distant metastasis rather than regional relapse, but not significantly. Prognosis from the initial treatment and PSA relapse was significantly poor when the PSA-DT was short. CONCLUSIONS: PSA elevated exponentially in the relapsed prostate cancer patients after the endocrine therapy. PSA-DT was distributed in a very wide range, and this value was considered to reflect the malignant potential and prognosis of the cancer. PSA-DT may be useful for determining the strategy after relapse.