Synthesis, properties, and intratumoral evaluation of mitoxantrone-loaded casein microspheres in Lewis lung carcinoma.

Smooth, round, uniform bovine casein microspheres of 1-5 and 10-20 microns size were readily prepared by a steric stabilization technique previously developed in this laboratory for synthesis of albumin microspheres. The avid phagocytic uptake of casein and albumin microspheres was demonstrated with fluorescein-labelled microspheres using a macrophage-like mouse myelomonocytic leukaemia cell line. Post-synthesis loading of 25% mitoxantrone was achieved for casein microspheres containing 20% polyglutamic acid. Preliminary intratumoural chemotherapy experiments with a mouse Lewis lung carcinoma indicated that mitoxantrone and mitoxantrone-loaded casein-polyglutamic acid microspheres exhibited lower toxicity when administered intratumorally.

Tolerance to long-term feeding of isolated peanut lectin in the rat: evidence for a trophic effect on the small intestines.

Previously we have shown that rats fed a diet containing raw peanut meal as the sole source of protein exhibited alterations in enzyme activity and composition of certain organs. To determine the effects of isolated peanut lectin on body growth and on the intestines, experiments were carried out in weanling, male, Sprague-Dawley rats fed a casein diet incorporated with purified peanut lectin at three levels, 0.004, 0.04, and 0.2% for 23 days. Body weight gain was normal with all three diets. In rats fed the 0.004 and 0.04% peanut lectin, there were no changes in any of the small intestinal mucosal parameters under study. However, in rats consuming the 0.2% peanut lectin diet, the proximal, mid, and distal third regions of the small intestines all showed marked increases in mucosal weight, protein, and DNA contents, but without altered villus morphology. Of the 3 brush border enzymes studied, namely maltase, gamma-glutamyltranspeptidase, and alkaline phosphatase, none was altered in activity in any region, suggesting that microvillus integrity was normal. These results are similar to the reported actions of red kidney bean lectin on the intestines. We conclude that peanut lectin at up to 0.2% of the diet does not inhibit food intake or growth of weanling rats and is apparently trophic for all areas of the small intestines.

Inhibition of tumor growth and metastasis by chronic intravenous infusion of prostaglandin E1.

The role of prostaglandins and their synthesis inhibitors in malignant disease is undefined. The following studies were done to determine the effects of continuous intravenous prostaglandin E1 (PGE1) or a prostaglandin synthesis inhibitor, indomethacin, on tumor growth and metastasis in mice bearing Lewis lung carcinoma. Male B6D2F1 mice underwent tumor implantation in the right axilla on day 0. After 10 days of tumor growth, mice underwent intravenous (IV) catheterization and were infused with either PGE1 at 3 micrograms/kg/minute (PG-LOW), PGE1 at 6 micrograms/kg/minute (PG-HIGH), indomethacin (INDO) at 1 microgram/kg/minute, or normal saline (NS). After 10 days of infusion, tumor volume, tumor weight, and the number of metastases greater than 2 mm in diameter were significantly decreased, and tumor doubling time was significantly prolonged in the PG-HIGH group compared to NS controls. None of the other experimental groups showed differences in these parameters. A second experiment with a similar experimental design was done infusing PGE1 at 6 micrograms/kg/minute and at 12 micrograms/kg/minute to determine the maximum dose response of IV PGE1. Again a decrease in tumor volume, tumor weight, and metastatic rates were identified when compared to saline control, but there were no significant difference between the two doses of PGE1.

Differential role of prostaglandin E1 on tumor metastasis.

Previous studies from our laboratory have shown that intravenous (iv) infusion of pharmacologic doses of prostaglandin E1 (PGE1) inhibits "spontaneous" metastases in mice bearing Lewis Lung carcinoma (LLC). This study was done to determine the effect of iv PGE1 on "artificial" metastases. Male hybrid mice (20 g) underwent iv catheterization and were begun on continuous infusions of either PGE1 at 10 micrograms/kg/min or 0.9% NaCl (NS). After 2 days, each mouse received an iv injection of 10(5) viable LLC cells and was continued on the infusion regimens. Pulmonary metastatic nodules were enumerated and measured 12 days after injection. The iv infusion of PGE1 increased the metastatic rate 6-fold, the number of metastases greater than or equal to 2 mm in diameter 7-fold, and the number of metastases greater than or equal to 3 mm in diameter 14-fold. Thus, PGE1 demonstrates differential effects on tumor metastases, acting as a promotor of artificial metastases, in contrast to its antimetastatic effect on an intact primary tumor. When the promotional effect of iv PGE1 on artificial metastases is evaluated in light of our previous studies, it can be concluded that the inhibitory effect of PGE1 on spontaneous metastases from an intact tumor is secondary to the effect of PGE1 at the level of the primary tumor, and not in the circulation or at the target organ of metastases (lung).

Correlation of estrogen, progesterone, and androgen receptors in breast cancer.

Breast cancer was induced in female Holtzman rats by intragastric administration of 7,12-dimethylbenz[a]antracene (DMBA). At tumor maturity, biopsies of viable tissue were obtained, frozen, and then assayed for estrogen, progesterone, and androgen receptor content. By simple linear regression analysis, progesterone receptor levels significantly correlated with both estrogen and androgen receptor levels, whereas estrogen and androgen receptor levels did not correlate with each other. Multiple regression analyses further substantiated the predictive value of the progesterone receptor for the other two hormone receptors. Knowledge of breast tumor androgen receptor levels may further enhance the value of the estrogen receptor and progesterone receptor in hormonal responsiveness. Further, the progesterone receptor may be the most sensitive of the steroid hormone receptors for selecting patients likely to respond to hormonal therapy.

Lability of steroid hormone receptors following devascularization of breast tumors.

Ischemia may invalidate hormone-receptor analyses. This study determined the effects of progressive ischemia on steroid hormone-receptor analyses. Breast cancer was induced in 50- to 60-day-old female Holtzman rats by intragastric administration of 25 mg of 7,12-dimethylbenz[a]anthracene. After 90 days, rats were anesthetized and breast tumors were devascularized in vivo. At 0, 30, 60, 90 and 150 minutes, a biopsy specimen from each tumor was taken and rapidly frozen. Steroid binding capacity for estrogen (ER), progesterone (PR), and androgen (AR) receptors was determined by incubation with tracer receptor ligand. Ischemia decreased ER and AR levels by 30 minutes, whereas PR levels were unchanged through 150 minutes of ischemia. Following mastectomy, tylectomy, or breast biopsy, PR may be the most reliable of the hormone receptors for determining endocrine-responsive breast cancer. However, for accurate determination of all hormone receptors, specimens should be frozen in liquid nitrogen immediately, then preserved at -70 degrees C, or processed immediately.

Partial hepatectomy decreases pancreatic parenchymal enzyme activity.

In previous studies of liver regeneration following partial hepatectomy (PH) in rats, we noted a decrease in serum amylase activity. The present study was carried out to determine if the decrease in serum amylase following PH corresponds to a decrease in pancreatic parenchymal exocrine enzyme activity. Male Wistar rats (275 g) underwent either 60% PH or sham laparotomy (SL). A third group, serving as controls (CON), received no surgical procedure. All groups were pair-fed a nutritionally balanced diet. After 48 hr, pancreas, blood, and liver were removed for analysis. Results showed that PH decreased pancreatic parenchymal protein by 43%, lipase by 44%, and trypsin by 22% compared to controls. Serum amylase decreased by 28%, a finding similar to previous experiments. However, pancreatic parenchymal amylase activity was not affected. It remains to be determined whether the reduced pancreatic function affects digestion and absorption.

Nutritional support by intraperitoneal dialysis in the rat: maintenance of body weight with normal liver and plasma chemistries.

A rat model was developed to study intraperitoneal (ip) dialysis as a means of total nutritional support. Rats (200 g) were implanted ip with a catheter device and connected to a rodent infusion assembly. An automated system exchanged 10-ml volumes of a 37 degrees C solution containing 10% dextrose, 2% amino acid solution plus micronutrients. Rats were adapted over 3 days to a schedule of 16 1-hr cycles/day, and continued on this regimen for another 4 days. Rats subjected to this program maintained similar body weight, nitrogen balance, plasma chemistries, and liver tests in comparison to control animals fed per os in isocaloric and isonitrogenous amounts. Efficiency of peritoneal absorption for both glucose and amino acid was 95%. Histological examination of intraabdominal organs revealed only mild inflammation. This model is applicable to studies involving nutritional support via the peritoneal cavity, a technique which may be of value in patients with sensitive fluid balances (cardiac, renal, or pulmonary failure).

Comparison of glucose and glucose plus lipid as caloric sources in parenterally fed rats.

This study investigated the etiology of fat infiltration of the liver during total parenteral nutrition. We measured the content of liver lipids, serum lipids, liver lipogenic enzymes, rates of in vivo fatty acid synthesis, and carcass composition in rats during continuous intravenous (iv) and intragastric (ig) feeding of two diets containing either 100% glucose or 75% glucose-25% lipid (20% Intralipid). Two groups of orally (O) fed rats were given solid diets similar to either the glucose or glucose-lipid solution in energy and nitrogen content. All six groups of rats (285-295 g) received 230 kcal X kg-1 X day -1 and 766 mg N X kg-1 X day-1. Total liver fat was greater after feeding the glucose diet ig rather than iv. However, feeding the glucose-lipid diet ig but not iv reduced the accumulation of liver fat by 49%. There were no differences in serum glucose concentrations among the three groups fed the glucose solution. Serum glucose concentrations in iv and O rats fed either diet were not significantly different; whereas feeding the glucose-lipid solution ig lowered serum glucose compared with the 100% glucose solution. Insulin concentrations were similar among all groups. The concentrations of serum triglycerides and cholesterol were higher in the groups fed the glucose-lipid diet. The activities of the liver lipogenic enzymes and rates of fatty acid synthesis were higher in iv- and ig-fed rats receiving the glucose diet compared with the glucose-lipid diet.(ABSTRACT TRUNCATED AT 250 WORDS)

Dosage form and formulation effects on the bioavailability of vitamin E, riboflavin, and vitamin B-6 from multivitamin preparations.

The effects of formulation factors and pharmaceutical dosage form on the bioavailability of RRR-alpha-tocopherol (d-alpha-tocopherol), riboflavin, and pyridoxine hydrochloride were studied after administration of two capsule formulations and a tablet to 12 normal humans. Absorption of RRR-alpha-tocopherol was increased from the Aquabiosorb soft elastic gelatin (SEG) capsule formulation compared with the modified standard-SEG capsule and the commercial tablet. There were no significant differences in bioavailability of riboflavin and pyridoxine hydrochloride between the SEG formulation and the tablet albeit a trend toward consistent absorption was seen from the SEG formulation. The modified-SEG formulation exhibited significantly lower bioavailability for these water-soluble vitamins. The enhanced bioavailability of vitamin E and the trend towards faster and more consistent absorption of riboflavin and vitamin B-6 from the SEG formulation may be related to the surfactant vehicle employed and the attendant wetting properties. The results also suggest ethnic differences in vitamin bioavailability.

Causative factors for decreased pulmonary metastasis in parenterally fed mice.

Total parenteral nutrition (TPN) with fat and/or glucose as the caloric source is associated with a decrease in pulmonary metastasis in mice bearing subcutaneously implanted Lewis lung carcinoma. Five groups of white mice bearing Lewis lung carcinoma were assigned to receive various isocaloric and isonitrogenous oral and parenteral feedings: TPN, utilizing all nonnitrogen energy from glucose; per os, utilizing all nonnitrogen calories from glucose; electrolyte, utilizing nonnitrogen calories provided from a balanced casein diet and receiving an isovolemic infusion of electrolytes in the same composition as the TPN formula; 1/4 normal saline, also consuming the casein diet and receiving an isovolemic infusion of 1/4 normal saline; and an oral casein control (CON) without infusion. Results showed that there were no significant differences in tumor volume changes or tumor doubling time among the groups. However, tumor weight was significantly lower in groups receiving the TPN solution either orally or parenterally in comparison to the oral casein control. Pulmonary metastases were significantly lower in all parenteral groups, irrespective of solution composition, compared to the CON group. Thus it appears that parenteral fluid load rather than composition of the solution is the causative factor for the decrease in pulmonary metastases.

Decreased lung metastasis and tumor growth in parenterally fed mice.

The influence of alternate forms of nutritional support on primary tumor growth rate, tumor DNA synthesis rate, and number of lung metastases was examined in Swiss mice bearing subcutaneously implanted Lewis lung carcinoma (LLC). From Day 14 through 22 postimplant, mice were fed by continuous intravenous infusion of dextrose/amino acid (TPN), were offered the same solution from a feeding bottle (PO), were offered a casein-based, solid diet (CASEIN), or were infused with an electrolyte (ELECT) solution while energy and nitrogen were provided from the casein diet. Tumor weight and doubling time were decreased in the PO group compared to CASEIN; however, host weight decreased by 22% in the PO group. Tumor weight and DNA synthesis were decreased in the TPN group compared to CASEIN, and host weight increased by 4.6%. The decreased rate of tumor growth in the PO group was not reflected in a decrease in DNA synthesis, perhaps a result of the circadian pattern of DNA synthesis as previously reported for LLC. The number of metastatic lung nodules was significantly decreased in both the TPN and ELECT groups compared to PO and CASEIN, suggesting that intravenous fluid load rather than nutrient intake was the causative factor. In this host-tumor system, parenteral feeding was associated with a decrease in primary tumor weight and DNA synthesis rate, maintenance of host weight, and a decrease in pulmonary metastatic disease compared to mice fed a conventional diet.

Total parenteral nutrition in mice bearing a metastatic carcinoma: tumor growth, metastasis and immunologic parameters.

The role of dietary manipulation of tumor growth, metastasis and immunologic parameters was studied in mice bearing Lewis lung carcinoma. Fourteen days following subcutaneous tumor implant, groups with tumor and their non-tumor bearing counterparts were assigned to one of the following feeding protocols: total parenteral nutrition (TPN), per oral (PO) intake of the parenteral diet, an oral casein diet (CAS), or electrolyte infusion plus the casein diet (ELECT). Intakes of energy and nitrogen were similar among all groups. Mice were killed 12 days later and peritoneal macrophages were tested for phagocytic activity. Tumor growth and metastasis were decreased from both infusion regimens with minimal loss of body weight as compared with casein fed mice. PO mice also showed lower tumor weight but metastasis was as great as in the casein group. Non-tumor-bearing infused mice showed depressed thymic weight, but thymic weight was not further reduced in tumor-bearing infused mice. PO feeding afforded no such protection in the presence of the carcinoma. Splenomegaly was observed in tumor-bearing mice on all regimens, but mice maintained on the parenteral diet demonstrated the largest proportion of macrophages containing nuclear debris. Analysis of free macrophages indicated no effect of diet regimen on non-immune phagocytic activity in both tumor-free and tumor-bearing mice. Possible alteration of splenic macrophage intracellular digestive capacity or phagocytic activity was suggested as a result of TPN.

Bacterial endotoxin retention by inline intravenous filters.

Filters used in i.v. administration sets were tested for their ability to retain bacterial endotoxins for up to 96 hours of continuous infusion. Inline filters composed of cellulose ester, polyacrylate, polypropylene, polyethylene, or Posidyne Nylon 66 were used during continuous infusion of 5% dextrose injection at 83 mL/hr. One milliliter of inoculum containing 10(8) Escherichia coli was injected through a port upstream from the filter. A bacterial filter was used to monitor the sterility of effluent from the inline filters. The effluent was tested with limulus amebocyte lysate (LAL) that could detect endotoxin concentrations greater than 50 pg/mL. A control solution was monitored for viability of the bacteria throughout the course of the study, and positive endotoxin controls were used to confirm the sensitivity of the LAL. Samples of effluent were tested at 0, 4, 19, 24, 48, 72, and 96 hours. Effluent from all filters was sterile throughout the study. LAL assay indicated that only the effluent from filters containing Posidyne Nylon 66 was free of endotoxins for 96 hours. Effluent from the other filters contained endotoxins immediately after injection of the E. coli. Of the inline filters tested, only the one composed of Posidyne Nylon 66 was able to retain E. coli endotoxin for 96 hours. Further study is needed with E. coli and other microorganisms that are likely contaminants of i.v. infusions.

Parenteral handling precaution: particulate introduction resulting from needle sheath contamination.

The purpose of this in vitro experiment was to determine if inadvertent needle cap nicking would introduce particulate contamination. After a preliminary observation (n = 21), a more thorough confirmatory experiment (n = 28) was performed. Twenty-four of 28 needles were contaminated before, and 18 of 28 needles after, entry into a Latex injection port; 5 of these 18 needles remained contaminated post sterile water injection and withdrawal from the port. Since 19 of 24 contaminated needles lost their contamination within the simulated injection system, it was proposed that inadvertent needle cap particulate contamination to needle shafts may be introduced into in vivo tissue or delivered intravascularly.

Cost containment using cysteine HCl acidification to increase calcium/phosphate solubility in hyperalimentation solutions.

The purpose of this study was to determine if (1) the calcium/phosphate insoluble product was inversely related to pH [when cysteine HC1 (CH) was added as neonatal supplementation at 0.5 mM/kg/day to hyperalimentation (HAL) solutions] and (2) the potential cost savings to the hospital. The pH of the HAL solutions was adjusted by adding various amounts of CH to the HAL solution. HAL solutions containing 27 mEq of calcium/liter and 30 mEq (15 mM) of phosphate/liter were compounded. Ten-milliliter aliquots were analyzed at 0, 12, 24, and 48 hr. All samples (n = 56) were filtered (0.22 mu), viewed with 7-10,000 X magnification scanning electron microscopy, and qualitatively analyzed with a Philips Energy Dispersive X-Ray Analysis System equipped with a SW9100 Microprocessor. Calcium/phosphate insoluble product was present in the 0-, 12-, 24-, and 48-hr samples from the CH-free solutions. The solutions containing 759 mg (4.17 mM)/liter of CH however, remained free of precipitant. This investigation demonstrated that addition of CH to HAL can foster significant cost containment (projected $82,000/yr tangible hospital savings) by the elimination of current calcium/phosphate separation procedures for neonates on parenteral nutrition.