Streptavidins with intersubunit crosslinks have enhanced stability.

Natural tetrameric streptavidin has two subunit interfaces; one is a strong interface between subunits in a tightly associated dimer, and the other is a weak interface between a pair of such dimers (dimer-dimer interface). To test whether strengthening the weak dimer-dimer interface could provide streptavidin with additional structural stability, covalent crosslinks were introduced between adjacent subunits through the dimer-dimer interface. Specific crosslinking sites were designed by site-directed mutations of His-127 residues that are in close proximity in natural streptavidin. The first and second streptavidin constructs have a disulfide bond and an irreversible covalent bond, respectively, between two Cys-127 residues across the dimer-dimer interface. The third variant is a hybrid tetramer consisting of two different streptavidin species, one having lysine and the other aspartic acid at position 127, which are covalently crosslinked. All streptavidin constructs with intersubunit crosslinks showed higher biotin-binding ability than natural core streptavidin after heat treatment. All of these crosslinked streptavidins retained bound biotin more stably than natural core streptavidin in guanidine hydrochloride at very acidic pH. These results suggest that the introduction of covalent bonds across the dimer-dimer interface enhances the overall stability of streptavidin.

Carcinogenic effects of diisopropanolinitrosamine in Sprague-Dawley rats.

The carcinogenic effects of diisopropanolinitrosamine (DIPN) were tested in Sprague-Dawley rats and were then compared with results produced earlier by the same substance in Syrian hamsters. In addition to the similarities, several differences were noted; for example, DIPN caused pancreatic tumors in all the hamsters, but only 1 pancreatic tumor was observed among the 150 rats. Administration of DIPN to rats led to the development of neoplasms in the nasal and paranasal cavities, lungs, thyroid gland, esophagus, liver, and kidneys. The highest tumor incidence in rats was in the nasal cavities. Almost all the pulmonary neoplasms were malignant and were usually squamous cell carcinomas. We found a 15.4-50% incidence of malignant tumors of the thyroid gland; we also noted that thyroid neoplasms occurred at almost the same rate (36.4-50%) in the groups given 1/5, 1/10, and 1/20 the median lethal dose.

Intrasplenic tumor formation in the syrian hamster (Mesocricetus auratus) after tracheal implants and treatment with diethylnitrosamine.

In two simultaneous experiments (A and B), tracheal tissue from 25 young Syrian hamsters was implanted in their mothers' spleens. In A, after tracheas from 3-week-old untreated donor offspring were implanted into their mothers, the mothers received 18 mg diethylnitrosamine (DEN)/kg body weight, sc, twice weekly for 20 weeks. Thereafter, the implants were removed and transferred to the spleens of other untreated offspring that were then similarly treated with DEN for 20 weeks. In B, the procedure was similar except that the donor offspring were pretreated with DEN for 20 weeks, beginning at the age of 8 weeks. The average treatment time for the implants in experiment A was 65 weeks; in B, 44 weeks. In A, only four implants demonstrated a papilloma or a papillary polyp. In B, such neoplasms developed after a much shorter treatment time, and in three cases, squamous cell carcinomas with invasive growth in to the splenic pulp were formed.

Carcinogenic effect of N-nitroso-2,6-dimethylmorpholine in Syrian golden hamsters.

N-Nitroso-2,6-dimethylmorpholine was intragastrically administered to Syrian golden hamsters at four different dose levels [74 mg/kg body wt = 1/5 median lethal dose (LD50); 37 mg/kg body wt = 1/10 LD50; 18 mg/kg body wt = 1/20 LD50; and 9 mg/kg body wt = 1/40 LD50]. Up to a 71% rate of pancreatic tumors was induced. These tumors were adenomas or adenocarcinomas of ductal origin. In addition, neoplasms developed in different percentages in the nasal cavities, larynx, trachea, lungs, liver, gallbladder, kidneys, and forestomach. The number of tumors induced was not significantly dose-dependent.

Carcinogenicity of subcutaneously injected N-nitrosoheptamethyleneimine in European hamsters.

Male and female European hamsters (45 of each sex) recieved sc injections once weekly for life of N-nitrosoheptamethyleneimine (NHMI) at one-fifth the median lethal dose (LD50) (females: 44 mg/kg body wt; males: 66 mg/kg body wt), one-tenth the LD50 (females: 22 mg/kg body wt; males: 33 mg/kg body wt), or one-twentieth the LD50 (females: 11 mg/kg body wt; males: 16.5 mg/kg body wt). Survival times for both males and females were dependent on the dose of NHMI. Pulmonary neoplasms were induced in almost all the treated animals. They were histologically diagnosed as adenocarcinomas, squamous cell carcinomas, and mixed cell carcinomas. In addition, nasal cavity tumors developed in all hamsters of all treatment groups; these were papillomas, squamous cell carcinomas, and a few adenocarcinomas. Only 1 tumor of the larynx and 1 tumor of the trachea were observed. Several papillomas and a few carcinomas were also detected in the forestomach. The results were discussed with reference to previous findings in rats and Syrian golden hamsters.

Effects of cigarette smoke on the bronchial epithelium of Syrian hamsters: ultrastructural studies.

The effects of two types of research cigarettes, which differed in their total smoke delivery and condensate, on the bronchial epithelium of Syrian hamsters were examined electronmicroscopically. The animals were exposed once a day for 5 days each week for 1 year to the total smoke of the cigarettes. Two groups and a control group were killed 1 day after the last exposure, and one group 12 months after the last exposure. The bronchial epithelia of all smoke-exposed animals were hyperplastic, and their ultrastructure showed invaginations, tilt of nuclear axes, an increase in the number and size of lysosomes and multivesiculated bodies, and increased numbers of enlarged intramitochondrial granules. Squamous metaplasia, rarely found, was present at the stage of filamentous bundle formation. The induced alterations were independent of the type of cigarette used. One year after smoking was terminated, the alterations had neither reversed nor advanced, as compared to those occurring 1 day after the last smoke exposure.

Transplacental effects of diethylnitrosamine in Syrian hamsters as related to different days of administration during pregnancy.

Female Syrian hamsters were given a single sc dose of 45 mg diethylnitrosamine (DEN)/kg body weight on 1 of the 15 days of pregnancy. In the offspring of females treated on 1 of the first 11 days of pregnancy, no respiratory tract tumors were found. The offspring of mothers given DEN on 1 of the last 4 days (12-15) of pregnancy developed respiratory tract neoplasms at a rate of up to 95%. A lower incidence of tumors in other organs seemed independent of the day of DEN treatment.

Effect of three retinoids on tracheal carcinogenesis with N-methyl-N-nitrosourea in hamsters.

Male Syrian golden hamsters received 12 weekly intratracheal exposures to 0.5% N-methyl-N-nitrosourea with a special catheter. Following exposures, animals were randomized into 4 groups of 63 hamsters and placed on diets of lab meal or meal with 120 mg 13-cis-retinoid acid (CRA)/kg, 327 mg ethyl retinamide (ER)/kg, or 343 mg N-(2-hydroxyethyl)retinamide (HR)/kg for 6 months at which time all hamsters were killed. The observed incidences of tracheal epithelial neoplasms (No. of animals with tumors/total No. of animals) were 10/63 (lab meal), 22/61 (CRA), 24/63 (ER), and 17/62 (HR). The incidence of carcinomas (No. of animals with tumors/total No. of animals) were 4/63 (lab meal), 12/61 (CRA), 12/63 (ER), and 11/62 (HR). The weight loss and mortality relative to those in the group fed the lab meal were significantly in the group fed HR but not in the other retinoid-treated groups.

Carcinogenic effects of Di-n-propylnitrosamine, beta-hydroxypropyl-n-propylnitrosamine, and methyl-n-propylnitrosamine on Sprague-Dawlay rats.

The carcinogenic effects of di-n-propylnitrosamine and its two related compounds, beta-hydroxypropyl-n-propylnitrosamine (HPPN) and methyl-n-propylnitrosamine (MPN), were tested in rats and compared. All three substances induced neoplasms in the nasal and paranasal cavities, esophagus, and liver. MPN treatment caused the highest tumor incidence in the esophagus, whereas HPPN induced the most hepatic tumors.

Carcinogenicity of N-nitrosodiethylamine in hibernating and nonhibernating European hamsters.

Hibernating European hamsters reacted differently to sc injections of N-nitrosodiethylamine (DEN) than did European hamsters that were not hibernating. Hibernating animals tolerated higher dose levels but developed fewer neoplasms. In contrast, hibernating males had more pulmonary tumors than did the respective nonhibernators. However, the hibernating females of the low dosage group developed fewer lung tumors. The survival times were longer for the male hibernators than for the male nonhibernators. The organ specificity of DEN, as well as the morphology and histogenesis of the neoplasms, showed no differences between the hibernating and nonhibernating groups.

Induction of pancreatic duct of carcinomas in the Syrian hamster with 2,6-dimethylnitrosomorpholine.

Pancreatic duct carcinomas were induced in Syrian hamsters by intragastric administration of 2,6-dimethylnitrosomorpholine. As early as 30 weeks after treatment began, tumors were macroscopically visible in the pancreas; the maximum incidence of such tumors reached 71% by the end of the experiment. They were ductal in origin, and some metastasized to the lung. They were identified histologically as adenomas and carcinomas and developed in different parts of the organ without demonstrating any preferred localization.

Renal carcinogenic and nephrotoxic effects of the flame retardant tris(2,3-dibromopropyl) phosphate in F344 rats and (C57BL/6N X C3H/HeN)F1 mice.

Tris(2,3-dibromopropyl) phosphate (TBP) was administered in the feed at one of two concentrations to groups of 55 male and 55 female inbred F344 rats and to 50 male and 50 female (C57BL/6N X C3H/HeN)F1 mice. The high and low dietary concentrations of TBP administered orally were 100 and 50 ppm for the rats, respectively, and 1,000 and 500 ppm for the mice, respectively. For each rodent type, 55 animals of each sex were used as contols. In both rodent types, renal epithelial tumors developed at incidences that were significant for male and female rats and mice that received the doses. These tumors included renal tubular cell adenomas and carcinomas that developed from the proximal convoluted tubular epithelium. Among female mice and rats, hyperplasia and/or dysplasia of the proximal convoluted tubular epithelium with or without cystic dilatation of the tubules and increase in the size of cell nuclei were dose dependent and recognized as preneoplastic and/or toxic lesions. The comparative histogenesis of renal tubular neoplasms was discussed.

Tumors of the heart and stomach induced in European hamsters by intravenous administration of N-methyl-N-nitrosourea.

One hundred twenty captured European hamsters (Cricetus cricetus L.) were inoculated iv with 1/5, 1/10, or 1/20 the median lethal dose of N-methyl-N-nitrosourea (MNU) once weekly for 18 weeks. MNU induced mainly sarcomas in the heart and squamous cell carcinomas and sarcomas in the stomach. The highest incidence of such tumors was demonstrated by the lowest-dosage group. They were more frequent in males than in females. The effect of MNU was also localized at the site of administration and resulted in squamous cell carcinomas of the oral cavity. The possible fibroblastic or neurogenic origin of the sarcomas was explored.

Successful chemotherapy of experimental neuroendocrine lung tumors in hamsters with an antagonist of Ca2+/calmodulin.

The chemotherapeutic effect of B859-35, the (-)-enantiomer of dihydropyrine 3-methyl-5-3-(4,4-diphenyl-1-piperidinyl)-propyl-1,4-dihydro-2,6-dimethy l-4- (3-nitrophenyl)-pyridine-3,5-dicarboxylate-hydrochloride (niguldipine), was tested on tumors induced in Syrian golden hamsters by N-nitrosodiethylamine (DEN). Peripheral pulmonary adenomas/adenocarcinomas were induced in hamsters maintained under ambient air conditions by multiple s.c. injections of DEN for 20 weeks. We have reproducibly shown that within this time interval lung adenomas develop in a significant number of the animals. The carcinogen treatment was discontinued at this point and one group of these hamsters was given B859-35 intragastrically 5 days/week for 20 weeks while the second group of such tumor-bearing hamsters were kept for an identical time interval without further treatment. Neuroendocrine lung tumors were induced in hamsters maintained in an atmosphere of 60% O2 by multiple s.c. injections of DEN for 8 weeks. We have reproducibly shown that within this short time interval neuroendocrine lung tumors develop in a significant number of the animals. The carcinogen treatment was discontinued at this point and the animals were returned to ambient air conditions. One group of these tumor-bearing hamsters was then given B859-35 intragastrically 5 days/week for 20 weeks while a second group of these hamsters was kept untreated for an identical time interval. A control group was given s.c. injections of saline for 20 weeks under ambient air conditions. A dramatic and selective anticarcinogenic effect of B859-35 was observed on the neuroendocrine lung tumors and nasal cavity tumors induced by DEN/hyperoxia while tumors of larynx/trachea were not affected. B859-35 had no effect on peripheral adenomas/adenocarcinomas, nasal cavity tumors, papillary polyps of larynx/trachea, or liver tumors induced by DEN under ambient air conditions.

The carcinogenic effect of dimethylnitrosamine on the Chinese hamster (Cricetulus griseus).

Three groups of Chinese hamsters (20 males, 20 females/group) received weekly subcutaneous injections for life of dimethylnitrosamine (DMN) at dose levels 0.20, 0.10 and 0.05 of the LD50. Eighty-two to 100% of the animals developed tumors of vascular origin. The neoplasms were primarily hepatic hemangioendotheliomas and their incidence was unrelated to dose or sex. Although animals in the low dose group received a total dose which was one-third that of the highest dose group, their survival times were only slightly longer.

Induction of renal pelvic tumours in Sprague-Dawley rats by di-isopropanolnitrosamine.

Transitional cell papillomas and carcinomas of the renal pelvis were observed in Sprague-Dawley rats after subcutaneous (s.c.) treatment with di-isopropranolnitrosamine (DIPN). These tumours occurred unilaterally and appeared as early as 24 weeks after the commencement of treatment. The tumours frequently filled the entire renal pelvis and compressed the kidney parenchyma. The malignant neoplasms demonstrated infiltrative growth into the adjacent medullary parenchyma.

Carcinogenic effect of di-isopropanolnitrosamine on the rat thyroid gland.

Up to a 50% rate of thyroid gland tumours was induced in Sprague-Dawley rats after s.c. administration of di-isopropanolnitrosamine (DIPN). The neoplasms occurred both unilaterally and bilaterally, and were identified histologically as adenomas and adenocarcinomas of follicular epithelium origin. In a few instances, adenomas were observed as early as 18 weeks after commencement of treatment. Most adenocarcinomas could be observed macroscopically, and frequently demonstrated infiltration into the surrounding muscles.

Carcinogenic effects of different nitroso-compounds in chinese hamsters: N-dibutylnitrosamine and N-nitrosomethylurea.

Three hundred and twenty Chinese hamsters (Cricetulus griseus) (CH) were subcutaneously (s.c.) treated with 1/5, 1/10 or 1/20 the mean lethal dose (LD50) of N-dibutylnitrosamine (DBN) or N-nitrosomethylurea (NMU). In several respects these two substances produced the same organotropy in the CH as in the Syrian golden (SGH) and European hamsters (EH). DBN prolonged lung tumours and neoplasms of the injection site. NMU induced tumours of the injection site only.

Carcinogenic effect of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) in European hamsters.

Laboratory-bred European hamsters received intragastric administrations of N-methyl-N'-nitro-N-nitrosoguanidine (MNNG) once weekly for 20 weeks. The animals showed mainly squamous cell papillomas and carcinomas of the fore-stomach. The tumour incidence was higher in males (80%) than in females (30%). The average tumour latency was comparatively short (25 weeks).

Lung tumors induced by chronic inhalation of 1,2-dibromo-3-chloropropane in B6C3F1 mice.

1,2-Dibromo-3-chloropropane (DBCP) was inhaled by B6C3F1 mice of each sex at concentrations of 0.6 or 3.0 ppm for 6 h/day, 5 days/week, for 103 weeks. Untreated controls consisted of 50 mice of each sex. All surviving animals were killed at 104 weeks. Survival in the high-dose groups of female mice was significantly shorter than that in the corresponding controls. Alveolar-bronchiolar adenomas or carcinomas occurred in both male and female mice with dose-related trends (P < 0.007) in males and (P < 0.002) in females. Papillary carcinomas in the lung occurred in low-dose female mice at an increased incidence compared with controls (P < 0.006). Many induced tumors were morphologically different than those in controls.