Burden and predictors of hypertension in India: results of SEEK (Screening and Early Evaluation of Kidney Disease) study.

BACKGROUND: Hypertension (HTN) is one of the major causes of cardiovascular morbidity and mortality. The objective of the study was to investigate the burden and predictors of HTN in India. METHODS: 6120 subjects participated in the Screening and Early Evaluation of Kidney disease (SEEK), a community-based screening program in 53 camps in 13 representative geographic locations in India. Of these, 5929 had recorded blood pressure (BP) measurements. Potential predictors of HTN were collected using a structured questionnaire for SEEK study. RESULTS: HTN was observed in 43.5% of our cohort. After adjusting for center variation (p < 0.0001), predictors of a higher prevalence of HTN were older age ≥ 40 years (p < 0.0001), BMI of ≥ 23 Kg/M2 (p < 0.0004), larger waist circumference (p < 0.0001), working in sedentary occupation (p < 0.0001), having diabetes mellitus (p < 0.0001), having proteinuria (p < 0.0016), and increased serum creatinine (p < 0.0001). High school/some college education (p = 0.0016), versus less than 9th grade education, was related with lower prevalence of HTN. Of note, proteinuria and CKD were observed in 19% and 23.5% of HTN subjects. About half (54%) of the hypertensive subjects were aware of their hypertension status. CONCLUSIONS: HTN was common in this cohort from India. Older age, BMI ≥ 23 Kg/M2, waist circumference, sedentary occupation, education less, diabetes mellitus, presence of proteinuria, and raised serum creatinine were significant predictors of hypertension. Our data suggest that HTN is a major public health problem in India with low awareness, and requires aggressive community-based screening and education to improve health.

Tumoral calcinosis: a case report.

Tumoral Calcinosis is a distinct clinical and histological entity that is characterized by large periarticular deposits of calcium resembling a neoplasm and is found foremost in the region of hip, shoulder and elbow. We report a case of Tumoral Calcinosis in a 25-year-old male who presented to us with gradually increasing swelling of right axilla, and both hips of nearly two years duration. It was diagnostic enigma for the treating surgeons but with the help of an astute pathologist we diagnosed this rare condition and successfully treated it surgically.

Pregnancy induced mammary tumor specific effector cells are present long after parturition in a breast cancer model in rats.

Pregnancy is known to provide protection against 7,12 dimethylbenz[a]anthracene-(DMBA) induced mammary carcinogenesis in rats. We observed in earlier studies that splenocytes of parous rats have significant cytotoxicity against mammary tumor cells both in vitro and in vivo. However, it remains to be established how long these cytolytic cells persist after parturition in parous host. The present study was designed using parous rats, 36 or more days after parturition. We observed that fresh splenocytes from these rats had low cytolytic activity against mammary tumor cells. However, when these cells were re-stimulated with irradiated mammary tumor cells in vitro, they had significantly higher cytotoxicity against mammary tumor cells. These studies show for the first time that pregnancy induced cytotoxic splenocytes are present long after parturition and they can be restimulated in vitro to enhance the cytotoxic effect.

Tumorigenesis of rat mammary epithelial cells by N-nitroso-N-methylurea in an in vitro system: characterization of the microtumors.

Chemical carcinogenesis is a lengthy process that involves the rather loosely defined stages of initiation, promotion, and progression. Several model systems of mammary carcinogenesis have been designed to elucidate the mechanisms of chemical carcinogenesis. Most of these systems have included animal models. While organ specific chemical carcinogenesis can be initiated in these systems, the subsequent stages of promotion and progression are difficult to study in detail. Investigations on in vitro carcinogenesis have shown transformation of mammalian cells in culture; the transformational event, however, is difficult to discern within the monolayer culture. We have recently reported the development of an in vitro carcinogenesis system that allows both the initiation as well as the progression of mammary cells in a collagen gel matrix culture system. The cells transformed by a chemical carcinogen develop into discernible microtumors within the three dimensions of a collagen gel culture. Isolation of these microtumors from the collagen gel and subsequent culture in monolayer has produced cells capable of colony formation in soft agar. The present study further characterizes these microtumors originated in vitro by analysis of cell growth kinetics versus parallel control cells. In addition, flow cytometric and cytogenetic studies have been performed to investigate the chromosomal stability of these cells. It was also observed that the microtumors, produced in vitro from mammary epithelial cells of an inbred strain of rats, show the ability to form tumors upon transplantation into the fat pad of syngeneic hosts.

In vitro carcinogenesis of mammary epithelial cells by N-nitroso-N-methylurea using a collagen gel matrix culture.

Carcinogenesis is a lengthy process which eventually culminates in the transformed phenotype, cancer. However, much remains to be defined about the process of transformation. In vivo models for the study of the carcinogenic process present limitations because it is not possible to detect the premalignant stages in the animals. An in vitro model, on the other hand, facilitates the study of the carcinogenic process because it enables one to dissect out the crucial events required for carcinogenesis to occur. As carcinogenesis is believed to be a multistep process; initiation, promotion, and progression, a multistep, in vitro system has been devised in our laboratory to mimic each of these stages. We have previously shown the formation of "microtumors" in collagen gels, induced by 7,12-dimethylbenz(a) anthracene. In the present study the direct acting water soluble, mammary carcinogen, N-nitroso-N-methylurea (NMU) was used for tumorigenesis of mammary epithelial cells in culture. Mammary epithelial cells from virgin Sprague-Dawley rats were propagated and exposed to single or multiple doses of NMU while growing as a monolayer in glass petri dishes (initiation). Initiation cells were then plated into a collagen gel matrix culture. Prolonged growth in the collagen gels afforded for the progression of the transformed cells into discernable microtumors in the three-dimensional matrix of the collagen. The morphology of these "tumors" was determined by histologic sections of the gels. Fewer, if any, such structures existed in the untreated gels.

Pregnancy-induced antitumor cytotoxicity of T cell-rich fraction against mammary adenocarcinoma cells in rats.

Our earlier observations indicated that splenocytes from parous rats have high cytotoxic activity against mammary tumor cells in vitro. It has also been observed that this cytotoxic activity of splenocytes from parous rats can be adoptively transferred to virgin rats of same age. The present investigation is an attempt to determine the cell type in spleens of parous rats that cause the cytotoxicity against mammary tumors. The spleens from both virgin and parous rats were removed aseptically and T and B cell-rich fractions were separated. 7,12-Dimethylbenz[a]anthracene-induced rat mammary tumor epithelial cells were used as targets and the T and B cell-rich fractions from either virgin or parous rats were used as effectors. The parous rats were divided into two groups according to the time period after parturition. Group I contained rats that were 5-13 days after delivery, while the rats in group II were 14 or more days post parturition. In vitro cytotoxicity was determined by incubating the tumor cells with spleen cells in the target/effector ratio of 1:3, 1:10 and 1:30 and using the standard 51Cr release assay. In all ratio groups of T cell-rich fractions particularly in group II, there was significantly higher cytotoxicity against mammary tumor cells. None of the B cell-rich fractions from parous rats were significantly more cytotoxic than those from virgin controls.

Pregnancy-induced cytotoxicity of splenocytes against mammary tumor cells in rats.

Our earlier observations indicate the possibility of involvement of a 'host factor' in pregnancy-induced protection against mammary carcinogenesis. The present investigation is an attempt to determine if this "host factor" is of immunological nature. Rat mammary tumors induced by 7,12-dimethylbenz[a]anthracene were used as target, and splenocytes from parous rats of the same strain were used as effector cells. The parous rats were divided into two groups according to the time period after parturition. Group 1 contained rats which were 5-13 days after delivery, group 2 had rats 14 or more days after parturition. In vitro cytotoxicity was determined by incubating the tumor cells with spleen cells in the target:effector ratios of 1:3, 1:10 and 1:30. Both groups 1 and 2 showed significant lysis of mammary tumor cells. These results were confirmed by the trypan blue exclusion test. The results showed a significantly higher cytotoxic capability of the spleen cells from parous rats against mammary tumor cells as compared to spleen cells from age-matched nulliparous rats.

Inhibition of mammary tumorigenesis in virgin rats by adoptive transfer of splenocytes from parous donors.

Splenocytes from parous rats have been previously found to have cytotoxic activity against mammary tumor cells in vitro. Experiments were carried out to determine if this pregnancy-induced cytotoxic nature of the splenocytes is inherent and transferable. Splenocytes from parous rats wer adoptively transferred to a group of virgin rats. Another group of age-matched, virgin rats received splenocytes from virgin donors in a similar way. After a period of rest, at the age of 55 days, the rats belonging to both of the groups, received 7,12-dimethylbenz(a)anthracene (DMBA) intragastrically. A third group of untreated virgin rats were also given the chemical carcinogen the same way as above and were considered as intact controls. The rats were monitored for development and growth of mammary tumor from 60 days of DMBA administration. After 4 months of DMBA administration the rats were sacrificed and mammary glands were examined for tumors. Mammary glands with no visible tumors were taken for whole mount preparation, to be examined for microscopic lesions. The results showed that 33 of 41 intact control rats, developed tumor and 27 of the 34 rats that received spleen cells from virgin rats developed tumors. Of the rats that received spleen cells from parous rats, only 18 out of 37 rats developed tumors, indicating an inhibition of tumor induction in these rats. Growth rate of the tumors in this group was also slower than in the control groups.

In vivo antitumor activity of tetrahydrobenz(a)anthraquinone derivatives.

Female rats bearing DMBA induced mammary carcinomas were treated with a new anthraquinone derivative related to mitoxantrone. The drug was administered at a dose of 5 mg/kg daily for 10 days. Tumor regression was noted in 78% of the animals with growth rate reduced from +74.91% to -12.60%. The results demonstrate that replacement of a polar hydroxy group of mitoxantrone with a nonpolar hydrocarbon moiety does not impair its antitumor activity and in fact may influence its availability to endocrine systems.

Collagen gel culture of rat mammary tumor cells as an assay system for determination of therapeutic efficacy of chemotherapeutic agents.

Collagen gel culture of rat mammary epithelial cells was used as an in vitro assay system for determination of the therapeutic efficacy of three cytotoxic agents commonly used in the treatment of human breast cancer, namely 5-fluorouracil (5-FU), methotrexate, and Adriamyin (ADR). The same three drugs were also evaluated in vivo, and a good correlation was obtained between the results in these two systems. A 9-d culture was shown to be more reliable than a 12-d culture, because nondrug-related cell mortality became a confounding factor after 12 d. Although further experiments are necessary, it is suggested that collagen gel culture may well prove to be a useful assay system for determination of sensitivity of tumor cells to cytotoxic drugs with possible clinical applications in the choice of treatment modality administered to cancer patients.

Inhibition of mammary carcinogenesis in rats by dietary restriction.

Mammary tumors were induced by 7,12-dimethylbenz[a]anthracene (DMBA) in Sprague-Dawley female rats kept under different dietary restrictions. Starting at 40 days of age, 4 groups of rats were either full-fed or fed 25%, 50% or 80% of their daily ration. At 55 days of age DMBA was given by intravenous injection. Rats were continued on the restricted diet until 150 days after carcinogen treatment. Rats on 25% diet lost weight rapidly and the experiment had to be terminated. Rats on the 50% diet maintained a lower body weight throughout the experiment; only 12% developed tumors. Rats on the 80% diet lost weight initially, but at the termination of the experiment, there was no significant difference in body weight between this group and the full-fed controls. Of the rats on 80% diet, 34% developed tumors, compared to 92% tumor incidence in the full-fed controls. Vaginal smears were normal in the animals fed the 80% diet, while some irregularity was observed in the 50% group. Breeding capability in rats on the 80% diet was not affected, since there was no observable difference in the pregnancy rate between these animals and their controls. There was also no difference in plasma level of estrogen between the 80% diet group and the full-fed controls at the time of carcinogen treatment. [3H]Thymidine labelling index was significantly affected by 50% restriction of diet while there was no significant change in the 80% group.

Prevention of mammary carcinogenesis in rats by pregnancy: effect of full-term and interrupted pregnancy.

In this study, the role of parity in conferring protection of the mammary gland against chemical carcinogenesis induced by 7,12-dimethylbenz(a)anthracene (DMBA) was investigated. Experiments were also carried out to determine if an 'interrupted' pregnancy was capable of reducing the incidence of mammary tumour induction. Since it has been suggested that morphological development or the proliferative pattern of the mammary gland at the time of carcinogen administration may be involved in reducing the susceptibility of the mammary gland to chemical carcinogenesis, experiments were designed to elucidate the possible influence of these two factors. Sprague-Dawley female rats were mated and were either allowed to complete pregnancy and parturition or were subjected to Caesarian section on day 5, 10 or 15 of the pregnancy. When DMBA was administered i.v. to animals which had been allowed to complete a full-term pregnancy, only 14% developed tumours, compared to 70% in age-matched nulliparous controls. Termination of the pregnancy on days 5, 10 or 15 was as effective in reducing tumour incidence as full-term gestation and parturition, but still resulted in partial and statistically significant inhibition, compared to age-matched nulliparous controls. There was no significant difference in 3H-thymidine labelling index (LI) at the time of DMBA treatment in the parous rats compared to age-matched nulliparous controls. We also observed no significant differences in the morphological development of the mammary gland in parous and nulliparous rats of the same age. These results indicate that the protective mechanism may not lie in the mammary gland per se, but may indeed be a host factor, such as hormonal or immunological changes occurring in the host as a result of the pregnancy.

Role of serum and hormones during the growth and development of rat mammary tumor epithelial cells in collagen gel culture.

The characteristics of hormone-dependent rat mammary tumors in response to serum and hormones were determined in collagen gel matrix culture. Epithelial cells from 7,12-dimethylbenz[a]anthracene (DMBA)-induced mammary adenocarcinomas were embedded in collagen gel and the effect of estrogen, progesterone, prolactin, insulin, and serum was tested. The total cell number and [3H]thymidine incorporation were used to determine the growth pattern of the cells in culture. It was found that in medium containing 20% porcine serum and supplemented with insulin, estrogen, progesterone and prolactin, both the cell number and [3H]thymidine labeling index increased with time, after an initial lag. Serum seemed to be essential to maintain growth of the tumor cells, because hormones alone, in the absence of serum, were unable to sustain growth of the cells. When estrogen, progesterone, prolactin, and insulin were tested individually in the presence of 20% porcine serum, only estrogen demonstrated a significant stimulatory effect.

Effect of estrogen and progesterone on cellular replication of human breast tumors.

A total of 54 infiltrating carcinomas of the breast were studied for estrogen receptor concentration in the tumor cytosol and thymidine-labeling indexes. The results showed that there was no significant difference in the level of thymidine-labeling index between 22 primary and 32 metastatic breast cancers. No significant association between the levels of thymidine-labeling index and the presence or absence of estrogen receptors was observed. The effect of "physiological" doses of estrogen and progesterone on cell proliferative activity was studied by the level of thymidine-labeling indexes in the tumor cells in 10 patients with multiple skin and s.c. metastases. Tumor biopsies were performed for labeling indexes both before and after hormonal treatment. The results showed that physiological doses of estrogen and progesterone induced a significant rise in thymidine-labeling index within 3 days after hormonal treatment in seven of the 10 tumors. Of the seven tumors that showed a rise in thymidine-labeling index, three were estrogen receptor positive and four were estrogen receptor negative. Of the three nonresponsive tumors, one was estrogen receptor positive and two were estrogen receptor negative. The study suggests that estrogen and progesterone can induce cell replication in both estrogen receptor-positive and -negative tumors.

Tumorigenesis of mammary gland by 7,12-dimethylbenz(a)anthracene during pregnancy: relationship with DNA synthesis.

The relationship between mammary cell proliferation during pregnancy and susceptibility to 7,12-dimethylbenz(a)anthracene (DMBA) was examined. DMBA was administered intravenously to Sprague-Dawley rats on the 5th, 10th or 15th day of pregnancy. [3H]thymidine labelling index (LI) of the mammary cells at the time of treatment with the carcinogen was determined and found to be higher in the pregnant rats than in age-matched virgin controls. In spite of the high proliferative index of the mammary cells, significant inhibition of tumorigenesis occurred in the pregnancy rats allowed to complete pregnancy and parturition following treatment with DMBA. However, when pregnancy was terminated by cesarian section shortly after treatment with DMBA, there was a significantly higher tumor incidence as compared to the "full-term" rats. It was observed that the earlier the pregnancy was terminated, the greater was the incidence of mammary tumors. This would indicate that the inhibitory effect of pregnancy is related to changes occurring during the later half of gestation. The differentiation of mammary cells for milk synthesis as pregnancy progresses is postulated to be a major reason for the observed refractoriness of the mammary cells to DMBA at that time.

Growth of mammary glands from monodispersed cells and susceptibility to 7,12-dimethylbenz[alpha]anthracene.

Monodispersed mammary cells inoculated into the subscapular fat pad of isologous female rats developed into full-formed ductal mammary glands. The growth pattern of these outgrowths followed that of normal mammary gland, as indicated by labeling index (LI). Tumorigenesis in these outgrowths induced by 7,12-dimethylbenz[alpha]anthracene (DMBA) was in direct proportion to the level of LI. A similar correlation of tumorigenesis and LI was also observed in the mammary gland of virgin female rats. Our data show that application of this technique in the study of mammary cell physiology and carcinogenesis would be very useful.

Enhancement of mammary tumorigenesis by dietary selenium deficiency in rats with a high polyunsaturated fat intake.

The effect of selenium depletion on mammary tumorigenesis following dimethylbenz[a]anthracene administration was examined in female Sprague-Dawley rats that were fed different levels and types of fats. Four basal diets deficient in selenium were used: (a) 1% corn oil; (b) 5% corn oil; (c) 25% corn oil; and (d) a high saturated fat diet containing 1% corn oil and 24% hydrogenated coconut oil. The comparable selenium-adequate diets were obtained by adding 0.1 ppm of selenium to each of the basal diets. In animals that received an adequate supplement of selenium, an increase in fat intake was accompanied by an increased tumor incidence when corn oil was used in the diets. A high saturated fat ration, on the other hand, was much less effective in this respect. Only in those rats that were maintained on a high polyunsaturated fat diet (25% corn oil) did selenium depletion result in a further increase in tumor incidence and tumor yield. Such an augmentation was not observed in animals given either a 1 or a 5% corn oil ration or a diet rich in saturated fat. Regardless of selenium status, almost all of the tumors found were adenocarcinomas. An enhancement of tumorigenesis as a result of selenium deficiency in rats fed the 1% corn oil regimen was detected provided a high dose of dimethylbenz[a]anthracene was used, suggesting that alterations in dimethylbenz[a]anthracene metabolism might be involved under this condition. The antioxidant property of selenium is discussed as a possible mechanism by which selenium protects against tumorigenesis, especially in animals with a high polyunsaturated fat intake.

Induction of mammary tumors in aging rats by 7,12-dimethylbenz[a]anthracene: role of DNA synthesis during carcinogenesis.

Two routes of administration were used to test the susceptibility of the mammary gland of the rat to 7,12-dimethylbenz[a]anthracene (DMBA) carcinogenesis in relation to age of the tissue. In one series of experiments, 60-, 70-, 90-, 120-, 150-, and 200-day-old female noninbred Sprague-Dawley rats were given DMBA iv. In parallel experiments, rats of the same ages as those above were given DMBA by local application. Mammary tumors developed in 80-90% of the 60- and 70-day-old rats and in 40% of the 90-day-old rats. Rats 120 days old and older were completely "refractory" to DMBA. In contrast, all rats, irrespective of their ages, developed tumors when DMBA was applied locally. The DNA synthesis labeling index (LI) of mammary glands of rats in two series of experiments was examined by [3H]thymidine incorporation and autoradiography at 24, 48, and 72 hours after DMBA treatment. DMBA, given iv significantly inhibited DNA synthesis in mammary glands, but DMBA applied locally significantly increased the LI of the mammary glands. Thus the LI of the mammary glands in 60-day-old rats increased from 8% (control, untreated) to 16% (rats receiving local applications of DMBA), whereas the LI of the mammary glands in 150-day-old rats increased from 1.15% (control, untreated) to 8% (rats receiving local application of DMBA).