DNA hypermethylation of extracellular matrix-related genes in human periodontal fibroblasts induced by stimulation for a prolonged period with lipopolysaccharide derived from Porphyromonas gingivalis.

OBJECTIVES AND BACKGROUND: The involvement of DNA methylation in periodontal disease is not clear. Lipopolysaccharide (LPS) derived from Porphyromonas gingivalis is involved in the progression of periodontal disease. We recently developed an in vitro model of LPS infection in human periodontal fibroblast cells (HPdLFs) for a prolonged period. In this study, we examined genome-wide analysis of DNA methylation in HPdLFs stimulated with LPS derived from P. gingivalis for a prolonged period. We noted the hypermethylation of extracellular matrix (ECM)-related genes and examined whether hypermethylation affected their transcription levels. MATERIAL AND METHODS: HPdLFs were grown in Dulbecco's modified Eagle's medium containing 10% fetal bovine serum. The culture was repeated, alternating 3 d with LPS derived from P. gingivalis and 3 d without LPS for 1 mo. Untreated samples were used as controls. DNA was analyzed using the human CpG island microarray. Quantitative methylation-specific polymerase chain reaction was carried out to confirm reproducibility of the microarray data. The expression levels of mRNA of the selected ECM-related genes from the data were analyzed by quantitative reverse transcription-polymerase chain reaction. RESULTS: We found 25 ECM-related genes with hypermethylation at the CpG island of the promoter region, which exhibited a fourfold greater hypermethylation than controls. Among these genes, hypermethylation of nine ECM-related genes, FANK1, COL4A1-A2, 12A1 and 15A1, LAMA5 and B1, MMP25, POMT1 and EMILIN3, induced a significantly downregulated expression of their mRNA. CONCLUSION: These results indicate that LPS derived from P. gingivalis may cause DNA hypermethylation of some ECM-related genes followed by downregulated expression of their transcriptional levels.

Predictors of delayed wound healing after endovascular therapy of isolated infrapopliteal lesions underlying critical limb ischemia in patients with high prevalence of diabetes mellitus and hemodialysis.

OBJECTIVES: Acceptable limb salvage rates underlie the widespread use of endovascular therapy (EVT) for patients with critical limb ischemia (CLI) secondary to isolated infrapopliteal lesions; however, post-EVT delayed wound healing remains a challenge. Predictors of delayed wound healing and their use in risk stratification of EVT in patients with CLI due to isolated infrapopliteal lesions are explored. METHODS: This was a retrospective multicenter study. 871 consecutive critically ischemic limbs were studied. There was tissue loss in 734 patients (age: 71 ± 10 years old; 71% male) who had undergone EVT between April 2004 and December 2012. The wound healing rate after EVT was estimated by the Kaplan-Meier method. The association between baseline characteristics and delayed wound healing was assessed by the Cox proportional hazard model. RESULTS: Diabetes mellitus and regular dialysis were present in 75% (553/734) and 64% (476/734) of patients, respectively; 67% of limbs (585/871) had Rutherford class 5 CLI; 8% (67/871) of wounds were located in the heel only; 25% (219/871) of limbs had Rutherford 6 (involving not only the heel); and 42% (354/871) of wounds were complicated by infection. The rate of freedom from major amputation at 1 year reached 88%, whereas the wound healing rate was 67%. Median time to wound healing was 146 days. By multivariate analysis, non-ambulatory status (hazard ratio [HR], 1.58; 95% confidence interval [CI] 1.31-1.91) serum albumin <3 g/dL (HR 1.42; 95% CI 1.08-1.86), Rutherford 6 (not only heel) (HR 1.68; 95% CI 1.33-2.14), wound infection (HR 1.24; 95% CI 1.03-1.50), EVT not based on angiosome concept (HR 1.28; 95% CI 1.06-1.55), and below the ankle (BTA) 0 vessel runoff after EVT (HR 1.45; 95% CI 1.14-1.86) were independent predictors of delayed wound healing. CONCLUSIONS: Non-ambulatory status, low albumin level, Rutherford 6 (not only heel), wound infection, indirect intervention, and poor BTA runoff were independent predictors for delayed wound healing after EVT in patients with CLI secondary to infrapopliteal lesions, and their use in risk stratification allows estimation of the wound healing rate.

Gelatinous transformation of the bone marrow in systemic lupus erythematosus.

Gelatinous transformation of the bone marrow is a rare disease characterised by a focal marrow hypoplasia, fat atrophy and accumulation of extracellular mucopolysaccharides abundant in hyaluronic acid, which is often associated with extreme malnutrition and weight loss. There are only two reports describing its association with systemic lupus erythematosus (SLE). One described underlying diseases in 155 cases of gelatinous transformation of the bone marrow and found one case with clinical diagnosis of SLE, but no clinical details were provided. The other described three SLE patients with gelatinous transformation of the bone marrow; however, two of these were cachectic and one was diagnosed with concomitant tuberculosis. We describe one active SLE patient without other comorbidities whose pancytopaenia was histologically confirmed as gelatinous transformation. The combination of high-dose steroid, intravenous immunoglobulin and mycophenolate mofetil improved the peripheral blood cytopaenia and reversed the bone marrow abnormalities.

The oligosaccharides in a recombinant hepatitis B virus surface antigen (HBsAg) carrying the pre-S2 region derived from yeast.

The N- and O-linked oligosaccharides in a yeast-derived HBsAg M protein (pre-S2 + S) were analyzed. Two major structures of the N-linked oligosaccharides bound to residue Asn4 were determined to be high-mannose type oligosaccharides, Man7GlcNAc2 and Man8GlcNAc2, by two-dimensional sugar mapping of the corresponding pyridylamino oligosaccharides. Peptide mapping of the M protein, sequence analysis of the glycopeptides after beta-elimination under reducing conditions and sugar-composition analysis revealed that the O-linked oligosaccharides were composed solely of mannose and bound to residue Ser5, Thr6, Thr7, Ser27, Ser28, Ser29 and Thr31 in the pre-S2 region.

A new model of plastic ankle foot orthosis (FAFO (II)) against spastic foot and genu recurvatum.

A plastic ankle foot orthosis (AFO) was developed, referred to as functional ankle foot orthosis Type 2 (FAFO (II)), which can deal with genu recurvatum and the severe spastic foot in walking. Clinical trials were successful for all varus and drop feet, and for most cases of genu recurvatum. Electromyogram studies showed that the FAFO (II) reduced the spasticity of gastrocnemius and hamstring muscles and activated the quadricep muscles. Gait analysis revealed a reduction of the knee angles in the stance phase on the affected side when using the FAFO (II). Mechanical stress tests showed excellent durability of the orthosis and demonstrated its effectiveness for controlling spasticity in comparison with other types of plastic AFOs.

[Carcinoma of the urachus: report of two cases].

Carcinoma of the urachus is a rare disease with poor prognosis. This is probably due to the difficulty of preoperative diagnosis. Radiological modalities have been considered to be of little value, but in many cases, preoperative diagnosis can be established with the aid of CT scan. In patients with urachal carcinoma, CT scan visualize the primary tumor and its extension beyond the bladder wall into the space of Retzius. Although about 150 cases of urachal tumor have been reported in the English literature, computerized tomographic findings have not been established. We present two cases and emphasize the usefulness of CT scan in preoperative diagnosis.

[A case of small simple renal cyst associated with renal cell carcinoma].

A 61-year-old male was admitted to our clinic for the purpose of further examination of the small renal mass discovered incidentally by ultrasonography. Subtraction angiography disclosed a small renal cell carcinoma of the upper-lateral margin of the left kidney dramatically. Enucleating surgery of the tumor was performed successfully (about 2 cm in diameter). Histological examination revealed a small simple renal cyst associated with renal cell carcinoma. The patient is well without evident metastasis 1 year postoperatively. Since this is a very rare case in the literature, we report this case and discuss the preoperative diagnosis of small renal cell carcinoma. We believe that ultrasonography is a simple and very useful method for mass screening of small urological malignancies.

[Familial vesicoureteral reflux].

Between August 1979 and January 1983, 43 cases of primary vesicoureteral reflux were treated in our hospital. Among them 9 cases of 4 families occurred familially. They were all siblings: 3 boys and 6 girls aged between 2 years and 10 years. Five cases except for 4 probands of each sibling were discovered after aggressive prospective family study, but they had no clinical symptoms that suspected vesicoureteral reflux; one of them was found to be in a preuremic condition. The other 2 siblings were examined by a prospective family study but demonstrated no vesicoureteral reflux. Therefore, vesicoureteral reflux could be demonstrated in 5 of the 7 siblings who had been examined by a prospective family study. This rate was significantly high (71%). Discussion is made about the necessity of prospective family study to discover asymptomatic patients with vesicoureteral reflux in the family, and the literature is reviewed.

Bilateral primary non-Hodgkin's lymphoma of the adrenal glands with adrenal insufficiency: a case report.

A 72-year-old woman with bilateral non-Hodgkin's lymphoma of the adrenal glands causing adrenal insufficiency is reported. The left-side lymphoma (diffuse large B-cell lymphoma) was removed surgically but the right-side lymphoma could not be removed. Complete response was obtained with subsequent combined chemotherapy and was maintained for 6 months with repeated chemotherapies. However, the patient died of liver recurrence 1 year after admission. In cases of malignant lymphoma forming a bulky mass, surgery with subsequent chemotherapy appears to reduce the cancer volume and improve the therapeutic outcome.

[Primary hyperparathyroidism: problems on surgical indication and procedure].

Between 1959 and Oct. 1990, 307 cases of primary hyperparathyroidism (PHPT) were operated on in our hospital. Among them, 23 cases (7.5%) were asymptomatic chemical type of PHPT, and the incidence of this type has been increasing these days. Various symptoms or signs including urolithiasis, bone disease, cardiovascular disease, gastrointestinal disease, diabetes mellitus and others were associated with PHPT. Especially, as a lethal factor, malignant tumors developed in 14 cases (4.6%); 9 cases of non-medullary thyroid cancer and tumors of other organs. In consideration of these associated disorders, the chemical type of PHPT should be operated prophylactically. In order to reduce operative complications, unilateral exploration is available for the cases of single normally localized adenoma; 85.7% of our 307 cases. Moreover, the positive rate of preoperative localized test by CT and ultrasonography for such adenomas is 78% in the recent 5 years. The predictive values of successful operation by unilateral exploration are 89% in the cases of normally localized single adenoma and 76% in all PHPT.

[The effect of kanpou medicine on the growth and aggregation of calcium oxalate crystals in vitro].

The effects of the compositional drugs of kanpou medicine on the growth and aggregation of calcium oxalate crystals were investigated in vitro. In the seeded crystal system, the inhibitory activities were calculated from the change of the number (Ia), total volume (Ig) and fractional volume (I) of crystals determined by a Coulter Multisizer. Among the tested drugs (Takusha, Akyou, Chorei, Bukuryou, Kasseki, Kinsensou, Kagosou), only Takusha and Kagosou had strong inhibitory activities on the growth and aggregation at the concentration of 5 micrograms/ml and 10 micrograms/ml (Ia greater than 80%, Ig, I greater than 90%). No significant correlation was observed between inhibitory activities and alcian blue precipitable polyanions (ABPP) of each drug. Thus some naturally existing substances composing the kanpou medicine may help prevent the stone formation in clinical use.

[The effect of takusha and kagosou on calcium oxalate renal stones in rats].

We examined the inhibitory effect of the two Kampo medicines, takusha and kagosou on the formation of calcium oxalate renal stones induced by ethylene glycol (EG) and 1 alpha(OH)D3 (1 alpha-D3) in rats. Wistar strain rats were divided into 4 groups (A: normal control, B: stone, C: kagosou, D: takusya). There was no significant difference in urinary calcium excretion or oxalate excretion between the stone group and kampo medicine groups. The calcium content of the kidneys was significantly lower in the takusha group than in the other two groups (Stone group and kagosou group). Takusha was effective in preventing oxalate stone formation in rats. Kagosou, which had strong inhibitory effect on calcium oxalate crystal growth and aggregation in vitro as well as takusha, was not effective against in vivo calcium oxalate stone formation in rats. These findings suggest that takusha prevents the formation of calcium oxalate stone by inhibiting calcium oxalate crystal growth and aggregation.

[Inhibitory effect of oral administration of Takusya on calcium oxalate crystallization in human whole urine].

We previously reported that Takusya had the inhibitory effect on in vitro calcium oxalate crystallization and in vivo stone formation in an animal model and it could be a prophylactic agent against calcium oxalate stone formation. We studied the effect of Takusya on calcium oxalate crystallization in human urine. Takusya (500 mg/day and 1,000 mg/day) was administered to 16 healthy men for 3 days and then 24-hour urine samples were collected to measure the urinary excretion of calcium, phosphate, magnesium, uric acid, creatinine, citric acid and oxalic acid. The urine samples before the administration of Takusya was used as a control. The size and distribution of crystals, which were formed in the urine samples by adding calcium chloride and sodium oxalate, were measured using the Coulter counter technique. Urinary magnesium excretion was significantly reduced by 1,000 mg/day of Takusya compared with the control (p < 0.05). The growth of crystals was significantly inhibited by 500 mg/day of Takusya in the large crystal formers whose urine created crystals more than 3.5 microns before the administration of Takusya (p < 0.05). These findings suggested that Takusya inhibited the growth of crystals formed in human urine.

[The effect of long-term alkali therapy for recurrent calcium stone patients with distal renal tubular acidosis].

Alkali therapy is widely accepted as a treatment for recurrent calcium stone patients with distal renal tubular acidosis (dRTA). Nine patients, five with complete and four with incomplete type of dRTA, were treated with alkali agents for more than three years; an average period of 10.8 years. In five patients, new stone formation and stone growth were completely prevented by this treatment. Among the four failed cases, three did not take enough alkali agents (in spite of our medication), and the other had recurrent urinary tract infection resulting in infectious stone formation. In conclusion, the long-term efficacy of alkali therapy for prevention of recurrent stone formation with dRTA is confirmed when the patient takes enough alkali agents and the urinary tract infection is well controlled.

[Extracorporeal shock wave lithotripsy for renal and ureteral calculi with MPL-9000. Correlation between stone burden and shock wave energy].

Clinical results of ESWL for renal and ureteral stone with MPL-9000 were presented. We also reviewed correlation between stone burden and shock wave energy. We studied 210 cases (295 sessions) of renal stone and 109 cases (154 sessions) of ureteral stone that were treated from November 1988 till January 1989 and from November 1989 till May 1990. Of those sessions, 76 sessions of renal stone cases and 53 of ureteral stone cases were carried out ambulatorily. Successful disintegration (within 4 mm) rate was 86% of renal stones, 84% of ureteral stones, and stone free rate after 6 months was 72% of renal stones and 84% of ureteral stones. As for the complications, brain infarction occurred in one case, subcapsular hematoma of the kidney in one, and arythmia in 5. No other severe side effects were observed. Between shock wave energy (the number of shock wave shots x voltage) and stone burden (long axis x short axis of the stone), a significant correlation was found in renal stone cases, but not in ureteral stone cases. E/B (shock wave energy/stone burden) was 33,100 (kv.shots/cm2) for the renal stone and 56,000 (kv.shots/cm2) for the ureteral stone. These results suggest that there is still another unknown factor, which is not related with renal stone, for successful disintegration of ureteral stone. We also confirmed that more energy is necessary for destruction of ureteral stone than for that of renal stone.(ABSTRACT TRUNCATED AT 250 WORDS)

[Evaluation of left ventricular early diastolic function by first pass radionuclide angiocardiography after dipyridamole infusion in patients with coronary artery disease--comparison with exercise test].

To evaluate the left ventricular early diastolic function after dipyridamole infusion or during exercise, the first pass radionuclide angiocardiography (FPRA) by a multicrystal gamma camera (SIM-400) was performed in 32 patients with coronary artery disease (CAD) and 14 normal subjects. Dipyridamole was administered intravenously (0.56 mg/kg in 4 min). The first third filling fraction (FF 1/3) and the first third mean filling rate (MFR 1/3) were calculated from the time-activity curve. FF 1/3 decreased and MFR 1/3 increased in both normal subjects and patients with CAD during exercise. After dipyridamole infusion, FF 1/3 and MFR 1/3 unchanged in normal subjects, in contrast, FF 1/3 and MFR 1/3 decreased in patients with CAD. Thus, dipyridamole infusion FPRA is more sensitive method to detect early diastolic dysfunction in patients with CAD than FPRA during exercise.

[Angiographic study of stenosis and calcification of coronary vessels in long-term dialysis patients: examination of risk factors for coronary calcification].

Coronary atherosclerosis and calcification score were assessed angiographically in 30 uremic patients receiving dialysis (8 patients with diabetes mellitus [DM dialysis group] and 22 patients without DM [non-DM dialysis group] who were suspected of having coronary artery disease because of their clinical symptoms and electrocardiographic findings. Thirty non-uremic subjects (11 with DM and 19 without DM) served as controls. Twelve dialysis patients (40%) did not have significant coronary artery disease, and the rate of significant coronary artery disease in dialysis patients overall was less than that in the control subjects. The calcification score of dialysis patients was significantly higher than that of the control patients (P < 0.0001), and also scores of the non-DM dialysis group and the DM dialysis group were higher than those of each group of control subjects. The scores were significantly correlated with the duration of dialysis in uremic patients as a group (r = 0.52; P < 0.01) and the non-DM dialysis group (r = 0.70; P < 0.01), but were not correlated with the duration of dialysis in the DM dialysis group. The scores were correlated with serum phosphate concentration, but not with serum calcium concentration, calcium phosphate product, alkaline phosphatase levels total cholesterol or age. Calcification scores were extremely high (> 21) in six uremic patients who had high serum c-terminal parathyroid hormone concentrations. These findings indicate that serum phosphate concentration, serum c-terminal parathyroid hormone concentration, and the duration of dialysis are closely associated with coronary calcification in long-term dialysis patients.

[Evaluation of changes in destructive spondylarthropathy (DSA) in long-term dialysis patients].

This study was undertaken to evaluate destructive spondylarthropathy (DSA) in patients long-term dialysis. The patients were investigated by plain X-ray examinations twice with a three-year interval, and the changes in severity of the illness in 66 patients undergoing dialysis for over 5 years were defined. Group A (9 patients) consisted of patients with progression in grades during the three years, while group B (57 patients) consisted of patients without deterioration. There was no differences in the two groups in age, duration and the modality of dialysis. However, in group A, the level of plasma beta 2-microglobulin (beta 2M) was 33.5 +/- 14.6 micrograms/ml at the first observation, and 33.1 +/- 15.1 micrograms/ml at the second (NS). In group B, the level of plasma beta 2M was 45.6 +/- 8.13 micrograms/ml at the first observation, and 39.7 +/- 9.18 micrograms/ml at the second (p <0.05). No improvement in DSA was observed. The result of this study suggests that DSA will not improve once it has become overt, and the management for lowering the level of beta 2M may prevent the occurrence and progression of DSA.

[Relative hypoparathyroidism associated with CAPD treatment using normo-calcemic (3.5mEq/1) dialysate: an approach from transperitoneal calcium balance].

We investigated factors affecting net transperitoneal calcium balance (Ca-BL) and the level of parathyroid hormone in relation to stepwise changes in serum calcium, by short PET (peritoneal equibrium test during 240 min: using 2000 ml of 2.5% dextrose dialysate containing 1.75 mmol/L Ca) in uremic patients undergoing stable CAPD. We calculated Ca-BL (mg/effluent/PET) of 244 effluents obtained from 90 patients receiving calcium carbonate as a phosphate binder without vitamin D supplementation. Their serum calcium level corrected with albumin (cSCa), alkaline phosphatase activity (ALP) and intact-PTH level was 9.7 +/- 0.9 mg/dl, 236 +/- 83 mIU/ml and 153.0 +/- 172.4 pg/ml, respectively. We proposed two statistic significant regression lines between Ca-BL and total drainage effluent volume (Ca-BL = 133X - 0.056: r = 0.981, P < 0.001), cSCa (Ca-BL = -12.9X + 123.6: r = 0.941, P < 0.01). Total drainage volume (TDV) and cSCa were two major factors affecting Ca-BL. A TDV level of 2430 ml/240 min-PET or more was required for positive Ca-BL in cases with 9.5-10.0 mg/dl of cSCa, using this linear regression analysis. A cSCa level of 9.6 mg/dl or more was also required for positive Ca-BL in cases with 2400-2600 ml/240min-PET. We also proposed a significant linear regression line between the intact-PTH level and cSCa (i-PTH = -90.5X + 1015.8, r = 0.973, P < 0.01). This line suggest that 200 pg/ml of intact PTH was obtained by 9.0 mg/dl or less of cSCa level in 90 CAPD uremic patients.(ABSTRACT TRUNCATED AT 250 WORDS)

[Cis-radiation therapy of locally advanced bladder cancer].

Locally advanced bladder cancer (clinical stage T2-T4) of 11 patients and ureteral cancer (clinical stage T3) of 3 patients was treated concurrently by CDDP and irradiation. CDDP, total 200 mg, was administered 20 mg/day X5 day on the 1st week and 4th week combined with 2 GY/day irradiation, total doses 40 GY. CDDP was given on one or 2 hours after irradiation on the day. The results were significantly improved that the tumor could be controlled by TUR, and the pathological specimens showed marked tumor necrosis or degeneration. In 63.6% of 11 invasive bladder cancers, bladder function was preserved without any evidence of tumor recurrence or metastasis during 15-37 months. From these results, it is concluded that cis-radiation therapy may preserve the bladder with locally invasive cancer.