Separation of urea, uric acid, creatine, and creatinine by micellar electrokinetic capillary chromatography with sodium cholate.

The capillary electrophoretic separation of the four nonprotein nitrogenous compounds (NPNs; urea, uric acid, creatine, and creatinine) typically employed in clinical and medical settings for the monitoring of renal function is described. Successful resolution of these compounds is achieved with the use of a bile salt micelle system composed of sodium cholate at phosphate buffer pH 7.4. The elution patterns of four NPNs are obtained within 30 min with a voltage of 30 kV. The effect of varying the applied voltage, temperature, and the mole ratio of phosphate buffer with bile salt surfactant on the migration behavior is also examined.

Discrimination of esophageal dysplasia with progression and nonprogression. High-resolution image analysis for surrogate end point biomarkers.

OBJECTIVE: To detect textural nuclear features correlated with nonprogression and progression in esophageal dysplasia. STUDY DESIGN: Asymptomatic adults from Heshun Commune, Linxian County, China were examined with a balloon sampler in 1983 fifty cases of moderate esophageal dysplasia and 68 cases of mild were selected for study. By means of an Axiomat microscope equipped with a TV camera, 100 visually normal intermediate squamous cell nuclei per specimen were randomly measured from routinely Papanicolaou-stained slides. RESULTS: Of 50 esophageal moderate dysplasia cases, 24 and 7 progressed to carcinoma within three and nine years, respectively. The other 19 cases remained stable or regressed to normal and were used as the control group. By means of chromatin features, correct specimen classification rates of 79.2% (19/24), 73.7% (14/19), 85.5% (6/7) and 84.2% (16/19) were achieved, respectively (P < .001). Of 68 cases classified as mild dysplasia, 16, 13 and 12 progressed to carcinoma within three, five and nine years, respectively. The other 27 cases remained stable or regressed to normal and were used as the control group. The correct specimen classification rates were 93.8% (15/16), 88.9% (24/27), 69.2% (9/13), 74.1% (20/27), 83.3% (10/12) and 77.8% (21/27), respectively, using chromatin features of the nuclei (P < .001). CONCLUSION: In this study, nuclear chromatin features measured by high-resolution image analysis could sufficiently well forecast the outcome of precancerous lesions and discriminate precancerous lesions with progression and nonprogression. It also can be employed as surrogate end point biomarkers in clinical chemoprevention trials. Stoichiometric staining and standard preparations should increase the correct classification rates in further studies.

Relevance of chromatin features in the progression of esophageal epithelial severe dysplasia.

Since 1983, a long-term clinical trial of esophageal carcinoma chemoprevention has been conducted in a high-risk area in China. From this study, 25 esophageal severe dysplasia patients without therapy were selected for analysis. After 5-year follow-ups, 14 cases progressed to esophageal carcinoma, while the other 11 cases remained stable. Three Papanicolaou's smears were used for each case, including one from the esophageal cytological examination at the beginning, two from the re-examinations three and five years later respectively. About 100 visually normal intermediate cells were randomly collected per slide by high resolution image analysis. More than 100 features (morphologic, densitometric, textural) were extracted. The classifications were made by means of stepwise linear discriminate analysis at the single cell level as on the specimen level using up to ten features. In all three comparisons of patients with progression and with regression at time of diagnosis, three years after diagnosis and five years later, the correct cell classification rates were about 70%. The subsequent specimen classifications by means of the a posteriori probability (APOP) distribution of the cells in each case led to 80% correct classification. All selected features reflected the chromatin structure of nuclei. The result demonstrated that the chromatin structures of esophageal epithelial cells in severely dysplasic patients are different between cases with and without progression. These results suggest the possibility of the application of image analysis in the clinical trials to find the dysplasia patients with higher risk of progression, in order to reduce the number of patients for therapy.

Segregation analysis of esophageal cancer in 221 high-risk Chinese families.

BACKGROUND: Until recently, environmental factors were considered of greatest importance in the etiology of esophageal cancer. Recent studies, however, have suggested that genetic factors also have a role. PURPOSE: Since no formal genetic study of this cancer has been previously reported, we carried out a statistical analysis to determine how important genetic factors are in the etiology of esophageal cancer in high-incidence areas of North China. METHODS: Using a logistic regressive model, we performed a segregation analysis on 221 high-risk nuclear families from the Yaocun Commune, Linxian, Henan Province of China, with at least one affected family member and with all offspring aged 40 years or older. Three models, the mendelian, the environmental, and the no-transmission models, were each compared with the general-transmission model that incorporated both genetic and environmental factors. RESULTS: According to Akaike's Information Criterion, the mendelian model provided the best fit for the data. By the chi-square test, the mendelian inheritance model was not rejected, but the environmental and the no-transmission models were both rejected. CONCLUSION: The segregation analysis indicated an autosomal recessive mendelian inheritance, with the alleged mendelian gene present at a frequency of 19%, causing 4% of this population to be predisposed to develop esophageal cancer. Large, unmeasured, residual familial factors, however, were also significant. IMPLICATIONS: Both an autosomal recessive gene and unexplained environmental factors appear to be important in the etiology of esophageal cancer in the subpopulation studied.

[Secondary prevention of esophageal cancer--intervention on precancerous lesions of the esophagus].

In 1983, intervention of precancerous lesion of esophagus was undertaken in the high risk area of esophageal cancer, Heshun Village, Linxian County. It had been expected that cancerous degeneration rate of esophageal dysplasia should be reduced by 50% so as the prevention of esophageal cancer could become possible. 6758 subjects of the general population aging from 40 to 65 were examined by esophageal exfoliative cytology, 1729 had marked dysplasia and 2411 had mild dysplasia of esophageal epithelium. Those with marked dysplasia were randomized into 3 groups to take their respective medication: antitumor B (Chinese herbs); retinamide (4-Ethoxycarbophenylretinamide) and placebo. The subjects with mild dysplasia were divided randomly into 2 groups for treatment by riboflavin and placebo. 95% of the subjects had taken 90% or more of the total medication for 3 years, at the end of which they were reexamined by esophageal exfoliative cytology. The reexamination rate was 94.1%. The incidence of esophageal cancer in the antitumor B group (3.9%) was reduced by 53% as compared with that of the placebo group (8.3%). This difference had statistical significant (means 2 = 7.672, P less than 0.05). The incidence of esophageal cancer in retinamide and riboflavin groups were reduced by 33.7% and 19% as compared with those of the control groups. The regression rate of dysplasia in the treatment groups were increased than that of the control groups. The above results showed that our hypothesis about the secondary prevention of esophageal cancer is correct. The intervention of precancerous lesion of the esophagus is effective in the prevention of esophageal cancer.(ABSTRACT TRUNCATED AT 250 WORDS)

[Carcinogenic and promoting effects of Roussin red methyl ester (RRME) on the forestomach epithelium of mice and esophageal epithelium of rats, and its inhibition by retinamide and vitamin C].

Carcinogenic and promoting effects of RRME as isolated from the pickled vegetables in Linxian County, a high incidence area of esophageal cancer, were studied in mice and rats. RRME alone did not cause tumor in the forestomach of mice and esophagus of rats. When the mice were intubated with a single dose of nitroso-sarcosine-ethylester (NSEE), the incidence of the forestomach carcinoma was only 9.5%. However, when the mice were given gastric doses of RRME after one single dose of NSEE, the incidence was increased to 41.0%. In rats, the tumor incidence was 5.3% in nitroso-methylbenzylamine (NMBzA) group, while in NMBzA kRME group, it was 20.7%. In rats intubated with NSEE for 7 times, no carcinoma appeared in esophageal epithelium; while followed by gastric doses of RRME, the incidence of esophagus carcinoma increased up to 63.2%. The experimental results show that RRME has distinct promoting effect on the process of cocarcinogenesis initiated by NSEE and NMBzA in the forestomach of mice and esophagus of rats, but without carcinogenic effect itself. Retinamide (RI) and massive dose of vitamin C showed an obviously inhibitory effect on promoting action of RRME in rats.(ABSTRACT TRUNCATED AT 250 WORDS)

[Carcinogenic and promoting effects of fish juice, preserved rice and salted dry fish on the forestomach epithelium of mice and esophageal epithelium of rats].

The carcinogenic and promoting effects of fish juice, preserved rice and salted dry fish from Nanau county, Guangdong province, a high incidence area of esophageal cancer, were studied in mice and rats. The homemade fish juice as well as fish juice in market, whether or not added with NaNO2, did not cause tumor in the forestomach of mice and the esophagus of rats. When the mice were intubated with an initiator, nitrososarcosinethylester (NSEE) twice, no carcinoma was found at the end of the experiment (D 120). Only papilloma appeared in the forestomach epithelium. The incidence was only 37.5%. However, when the mice were intubated with NSEE for 2 times followed by gastric doses of homemade fish juice, the tumor incidence in the forestomach was increased to 89.7%, in which 20.5% was carcinoma. The tumor and carcinoma incidences of initiator (NSEE and NMBzA) group and initiator + market fish juice group in mice and rats were without significant difference. The experimental results show that the homemade fish juice proved distinct promoting effect on the process of cocarcinogenesis initiated by NSEE in the forestomach of mice, while the market fish juice has no significant promoting effect on the forestomach epithelium of mice and the esophageal epithelium of rats. NSEE induced 31.6% carcinoma in the forestomach epithelium of mice on standard diet. While in mice fed with preserved rice and salted dry fish, the carcinoma incidence was increased to 63.6%. It appears that preserved rice and salted dry fish have promoting effect on the process of cocarcinogenesis initiated by NSEE in the forestomach of mice.(ABSTRACT TRUNCATED AT 250 WORDS)

[Photodynamic therapy (PDT) of bladder cancer in the rat].

PDT of rat bladder cancer, induced by N-butyl-N-(4-hydroxybutyl) nitrosamine (BBN) was studied. All animals were divided at random into three groups. Group 2 and 3 were treated by hematoporphyrin derivative (HPD)-laser PDT while group 1, without treatment, served as control. The results showed that the malignant lesions could be selectively and obviously destroyed, if the whole tumor area were sufficiently exposed to the laser irradiation. However, the normal bladder epithelium and muscle layer showed no histologic change. Similar reactions were found in papilloma of bladder which was considered as precancerous lesion. Thus, PDT may be beneficial to cancer prevention. Its role in prevention and treatment of bladder cancer should be further studied experimentally and clinically.

Metabolism of, and DNA methylation by, N-nitrosomethylbenzylamine in chicken.

The metabolism of 14C-nitrosomethylbenzylamine (NMBA) was studied in chicken. Following a single IV dose of 2 mg/kg, 14C-NMBA was cleared from the blood with a half-life of 3.8 min. At 10 min after administration 14C-NMBA was totally metabolized in the liver, whereas in the esophagus no measurable metabolic degradation had taken place. Maximum exhalation of radioactive CO2 occurred 1 h after IV administration of NMBA, and 11% of the total radioactivity had been exhaled as CO2 by 8 h. These results are compared with data on the metabolism of NMBA in the rat. The analysis of methylated bases in the DNA of different organs of chicken revealed that 7-me guanine was formed in all organs. The highest amount of 0(6)-me guanine was found in liver DNA, followed by kidney DNA. O6-me guanine was not detectable in any other organ. The O6-/7-me guanine ratio in DNA was calculated to be 0.05 and 0.02 for liver and 0.01 for kidneys.

Mutagenicity testing in the Salmonella typhimurium assay of phenolic compounds and phenolic fractions obtained from smokehouse smoke condensates.

Smokehouse smoke, which is used for flavouring meat products, was investigated for its mutagenic activity in the Salmonella typhimurium assay. We were chiefly concerned with the fractions free of polycyclic aromatic hydrocarbons but containing phenol compounds, which are responsible for the preservative and aromatizing properties of the smoke. The most abundantly occurring phenol compounds (phenol, cresols, 2,4-dimethylphenol, brenzcatechine, syringol, eugenol, vanilline and guaiacol) gave negative results when they were tested for mutagenicity at five concentrations up to 5000 micrograms/plate, with and without S-9 mix, using five strains of S. typhimurium. Even when phenol was further investigated in a variety of test conditions, no induction of his+ revertants was observed. When smokehouse smoke was condensed and fractionated the majority of the various phenolic fractions also gave negative results when tested at five concentrations using five strains of S. typhimurium. However there was a slight increase in the number of revertants in a few cases. The presence in the phenolic fractions of very small amounts of mutagenic impurities, the nature of which needs further investigation, cannot be excluded. These results support the further development of non-hazardous smoke-aroma preparations, based on the phenolic components of smokehouse smoke.

Examination of newly synthesized 2-chloroethylnitrosoureas in preterminal leukemia L 5222 and in two transplanted neurogenic tumors.

The chemotherapeutic activities of 11 chloroethylnitrosoureas, among them 10 newly synthesized compounds, were investigated in rat leukemia L 5222 and in two neurogenic rat tumors. 1-(4-Amino-2-methyl-5-pyrimidinyl)-methyl-3-(2-chloroethyl)-3-nitrosourea hydrochloride (ACNU, compound 1) and cyclophosphamide (12) served as reference substances. The newly synthesized compounds were 1-(2-chloroethyl)-1-nitroso-3-(2-carboxyethyl)-urea (2-carboxyethyl-CNU, 2), 1-(2-chloroethyl)-1-nitroso-4-methyl-4-formyl-semicarbazide (methyl-formyl-amino-CNU, 3), 1-(2-chloroethyl)-1-nitroso-3-(methylene-2-pyridyl)-urea(methylene-2-pyridyl- CNU, 4), 1-(2-chloroethyl)-1-nitroso-3-(methylene-2-pyridyl)-urea hydrochloride (methylene-2-pyridyl-CNU . HCl, 5) 1-(2-chloroethyl)-1-nitroso-3-(methylene-4-pyridyl)-urea hydrochloride (methylene-4-pyridyl-CNU . HCl, 6), 1-(2-chloroethyl)-1-nitroso-3-(3-pyridino)-urea (pyridyl-3-CNU, 7), 1-(2-chloroethyl)-1-nitroso-3-(3-pyridyl)-urea hydrochloride (pyridyl-3-CNU . HCl, 8), 1-(2-chloroethyl)-1-nitroso-3-[4(2,6-dimethyl-morpholino)]-urea (dimethyl-morpholino-CNU, 9), 1-(2-chloroethyl)-1-nitrosocarbamoyl-morpholine (chloroethyl-nitroso-carbamoyl-morpholine, 10), 1-(2-chloroethyl)-1-nitroso-carbamoyl-2,6-dimethyl-morpholine (chloroethyl-nitroso-carbamoyl-dimethylmorpholine, 11). Against both neurogenic tumors cyclophosphamide was distinctly superior to all nitrosoureas. In leukemia L 5222 all nitrosoureas except compounds 7, 8, 11 effected cures. Remarkable differences in toxicity could be observed between the nitroso compounds investigated.

Carcinogenic activity in rats of combined treatment with cyclophosphamide, methotrexate and 5-fluorouracil.

A total of 240 outbred Sprague-Dawley rats were treated with 3 different doses of the cyclophosphamide-methotrexate-5-fluorouracil (CMF) regimen adopted from clinical chemotherapy studies in breast cancer patients. Eighty untreated rats served as controls. Individual and total doses of the drugs applied were lower than corresponding doses used in human adjuvant therapy protocols compared on a mg/m2 basis. Lifelong observation of the animals demonstrated a strong dose-related carcinogenic response to the tested scheme. Main target organs of treatment-related neoplasms were the nervous system, the hematopoietic and lymphatic tissue, the urinary bladder, and the suprarenal gland. It is concluded that the CMF drug combination evokes carcinogenic responses in several organ systems in the rat and should be regarded as representing a carcinogenic risk to humans. Uncritical clinical use of the three-drug protocol should be avoided.