Poly-ε-Caprolactone Microsphere Polymers Containing Usnic Acid: Acute Toxicity and Anti-Inflammatory Activity.

Usnic acid (UA) has been studied by its pharmacological properties; however, it presents moderate toxicity, low solubility, and absorption by biological membranes. The aim of this study was to develop poly-ε-caprolactone microsphere polymers containing UA (UA-micro) and evaluate their acute toxicity and anti-inflammatory activity. The microspheres were prepared by multiple emulsion technique (water/oil/water) and characterized by the encapsulation efficiency, particle size, polydispersity index, and zeta potential. The acute toxicity of UA and UA-micro (25-50 mg/kg; p.o.) was evaluated in mice. The anti-inflammatory activity of UA and UA-micro was evaluated by subcutaneous air pouch and carrageenan-induced paw edema in rat, with measurement of inflammatory cytokines and MPO levels. The UA presented encapsulation efficiency of 97.72%, particle size of 13.54 micrometers, polydispersity index of 2.36, and zeta potential of 44.5 ± 2.95 mV. The UA-micro presented lower acute toxicity (LD50 value up to 2000 mg/kg; p.o.) when compared to UA. UA-micro and UA (25 mg/kg) significantly reduced paw volume and decreased MPO levels, whereas only UA-micro (50 mg/kg) reduced significantly IL-1ß, TNF-α, and NO levels in inflammatory exudate. These results suggest that controlled release systems, as microspheres, can be a promising alternative to reduce the toxicity of UA, making it a viable compound for inflammation therapy.

Developing a dynamic pharmacophore model for HIV-1 integrase.

We present the first receptor-based pharmacophore model for HIV-1 integrase. The development of "dynamic" pharmacophore models is a new method that accounts for the inherent flexibility of the active site and aims to reduce the entropic penalties associated with binding a ligand. Furthermore, this new drug discovery method overcomes the limitation of an incomplete crystal structure of the target protein. A molecular dynamics (MD) simulation describes the flexibility of the uncomplexed protein. Many conformational models of the protein are saved from the MD simulations and used in a series of multi-unit search for interacting conformers (MUSIC) simulations. MUSIC is a multiple-copy minimization method, available in the BOSS program; it is used to determine binding regions for probe molecules containing functional groups that complement the active site. All protein conformations from the MD are overlaid, and conserved binding regions for the probe molecules are identified. Those conserved binding regions define the dynamic pharmacophore model. Here, the dynamic model is compared to known inhibitors of the integrase as well as a three-point, ligand-based pharmacophore model from the literature. Also, a "static" pharmacophore model was determined in the standard fashion, using a single crystal structure. Inhibitors thought to bind in the active site of HIV-1 integrase fit the dynamic model but not the static model. Finally, we have identified a set of compounds from the Available Chemicals Directory that fit the dynamic pharmacophore model, and experimental testing of the compounds has confirmed several new inhibitors.

Barium enema: detection of colonic lesions in a community population.

OBJECTIVE: The purposes of this study were to assess the prevalence of colonic lesions detected at barium enema in a community practice, to compare the findings at barium enema between patients who are asymptomatic and have no known risk factors for colorectal cancer (screening group) and patients who have symptoms of colonic disease or have known risk factors, and to determine if a questionnaire about symptoms and risk factors is an appropriate screening tool. SUBJECTS AND METHODS: A self-administered questionnaire about colorectal symptoms and risk factors was given to 1779 patients scheduled for barium enema examination. On the basis of their responses, patients were divided into three groups: screening group (asymptomatic, without risk factors), symptomatic, and asymptomatic with risk factors. Each patient underwent a fluoroscopic barium enema. We then compared the results (number, histologic type, size of lesion(s), location in the colon, and Patient's age) and risk factors among the three groups. RESULTS: At least one lesion within the colorectum was found in 166 (9%) of 1779 patients at combined proctosigmoidoscopy and barium enema. The prevalence of lesions in the 111 patients with at least one lesion above the rectum at barium enema was 4% (32 of 738) for the screening group, 8% (38 of 476) for asymptomatic patients with risk factors, and 7% (41 of 565) for symptomatic patients (p = .015 when comparing the prevalence in the screening group with the prevalences in the other two groups). Twenty-nine percent of all colonic lesions were found in the screening group. Among the asymptomatic patients, risk factors that included a history of colorectal polyps and advanced age were associated with a significantly higher prevalence of colonic polyps found at barium enema. In the symptomatic group, if patients with histories of polyps were excluded, we were unable to identify other risk factors that led to a significantly higher prevalence of polyps. CONCLUSION: Asymptomatic patients without known risk factors have a significantly lower prevalence of colonic polyps than either symptomatic patients or patients with risk factors alone. Despite this lower prevalence, 29% of all lesions in our series were in the screening group. Assessment of risk factors through a patient questionnaire was not helpful in identifying a group of patients with a higher prevalence of lesions--except for a history of polyps. Management decisions based on a patient questionnaire should be approached with caution. When low-risk patients are denied screening examinations, a significant number of lesions will be missed.

Metachronous colon cancer in persons who have had a large adenomatous polyp.

OBJECTIVE: To determine, among persons who have had a large colon polyp, the risk of subsequent colon cancer at a site distant from that polyp. METHODS: Follow-up was done for 226 persons at the Mayo Clinic who had had a > or = 1-cm polyp demonstrated on barium enema between 1965 and 1970 and for whom yearly colon surveillance examination was recommended. Information was collected from Mayo Clinic records and from contact with patients, physicians, and other hospitals regarding the results of surveillance examinations and the development of colon cancer. Colon surveillance was routinely done at the Mayo Clinic using the technique of single contrast barium enema with vigorous manual fluoroscopic examination and proctoscopy. The expected rate of colorectal cancer (CRC) was calculated based on previously published rates for this community. RESULTS: Patients received, on average, four colon examinations in addition to the examination that discovered the index polyp. During 2126 person-years of follow-up, 16 persons developed a colon cancer at a location other than the site of the index polyp, in comparison with 4.0 expected cases, for a standardized incidence ratio of 4.0 (95% CI,2.3, 6.4). The cancers were large (mean 4.5cm) at presentation, and eight of the 16 cancers had been preceded within 3 yr by at least one negative barium enema. CONCLUSIONS: The rate to develop colon cancer in persons who have had a large colon polyp es about 4 times the expected rate, suggesting that such persons should be considered for aggressive colonoscopic surveillance. The failure to detect early cancer or its precursors by surveillance barium enema is probably explained by inherent insensitivity of single contrast barium enema.

Evidence for an intracerebral action of phenylalanine in stimulation of prolactin secretion: interaction of large neutral amino acids.

Phenylalanine stimulates PRL secretion when given orally or iv to normal individuals. To differentiate between hypothalamic and pituitary sites of action, we examined the effects of concurrent infusion of valine on the PRL response to iv phenylalanine in eight normal men. Since large neutral amino acids share the same high affinity blood-brain barrier transport system, entry of phenylalanine into the brain will be diminished by the simultaneous presence of high serum concentrations of valine; the pituitary, lying outside the blood-brain barrier, is not subject to these competitive effects. Valine significantly blunted the PRL response to phenylalanine, supporting an effect of phenylalanine within the blood-brain barrier to stimulate PRL release.

Aspartame and its constituent amino acids: effects on prolactin, cortisol, growth hormone, insulin, and glucose in normal humans.

Because large doses of phenylalanine stimulate prolactin secretion in man, we studied the acute effects of oral doses of aspartame (0.534 g, equivalent to the amount of aspartame in approximately 1 L beverage), aspartic acid (0.242 g), and phenylalanine (0.3 and 1.0 g) on serum prolactin and other hormones in normal humans. Prolactin was not stimulated by any of the aspartame meals, aspartic acid, or 0.3 g phenylalanine; a small rise in serum prolactin, similar to that produced by a high-protein mixed meal, followed ingestion of 1.0 g phenylalanine. Serum growth hormone showed no statistically significant changes in response to any of the experimental meals whereas cortisol and insulin fell slightly and glucose rose slightly during each of the meals. We conclude that these doses of aspartame do not alter secretion of prolactin, cortisol, growth hormone, or insulin in normal individuals.

Radiographic appearance of mucosal dysplasia associated with ulcerative colitis.

The radiographic appearance of dysplasia, a precancerous histologic change that is frequently present in colitic colons at high risk for cancer, was analyzed. Over a 5-year period, 170 patients with long-standing ulcerative colitis participated in a cancer surveillance program at our institution. On entry, all patients underwent radiography with a double-contrast barium enema and colonoscopy with multiple random biopsies. All radiographs were reviewed retrospectively and were correlated with the colonoscopic and histologic findings. Dysplasia, ranging from mild to severe, was found in the biopsy specimens from 26 (15%) of these patients (in 59 different regions). Thirteen of the 26 patients with dysplasia had colectomies, and the specimens were available for review. Most dysplasia is not radiographically visible, as was the case in 40 (68%) of the 59 regions found histologically to have dysplasia. In the 19 regions in which dysplasia was visible radiographically, it appeared as a solitary nodule or as several separate nodules in 11 (19%) of the 59 regions or as a close grouping of multiple adjacent nodules with apposed, flattened edges in 8 (14%) of the 59 regions. The radiographic finding of a nodule or of several separate nodules was not specific because these nodules were indistinguishable from the inflammatory nodules often present in colitic colons. On the other hand, the radiographic finding of a close grouping of adjacent nodules with apposed, flattened edges was associated with dysplasia 50% of the time. This radiographic appearance was seen in five of our 26 patients who had dysplasia. Our study shows that barium examinations are useful in some patients with chronic ulcerative colitis by suggesting the presence of dysplasia and directing the endoscopist to specific locations for biopsy.

Is there a safe interval between diagnostic invasive procedure and the barium enema study of the colorectum?

In an attempt to define the optimal interval of safety when a barium enema study of the colon follows a planned injury of the bowel by local treatment or biopsy (or both), we studied a group of 833 patients who were seen at the Mayo Clinic during 1978. In the study group, 886 polyps were destroyed by fulguration, 258 lesions were both sampled and fulgurated, and 126 areas were sampled for biopsy study. One hundred ninety-four patients had multiple lesions; in 193 of these, 2-18 diminutive polyps were fulgurated. Of the 846 barium studies in the 833 patients, 5 were done on the same day as the planned injury, 543 within 24 hours, and 174 within 72 hours. Four patients demonstrated extravasation of barium, but none had signs or symptoms of acute perforation nor did the resected surgical specimen demonstrate communication with site of injury.

Thyrotropin-releasing hormone levels decrease in hypothalamus of aging rats.

Because hypothalamic and extrahypothalamic levels of thyrotropin-releasing hormone immunoreactivity (TRH-IR) undergo profound changes during the prenatal and early postnatal period in rats, similar effects with advanced aging were anticipated. For this reason we measured hypothalamic and reproductive tissue levels of TRH-IR, hypothalamic levels of somatostatin (SRIF), and beta-endorphin (EP), serum levels of prolactin (Prl), growth hormone (GH), thyrotropin (TSH), and thyroxine (T4) in young, sexually mature and 24-28 month-old male Long-Evans and Sprague-Dawley rats. Hypothalamic and prostatic levels of TRH-IR were consistently reduced as were the levels of T4 in old rats compared to young controls. Aging did not change the ratio of TRH to the major TRH-like peptide in prostates, as determined by high pressure liquid chromatography (HPLC) or the levels of hypothalamic SRIF and EP. All of the hypothalamic TRH-IR in both old and young male rats consisted of TRH by HPLC. Falling hypothalamic TRH levels and TRH secretory capacity may play a role in the blunted TSH response to cold stress in old rats.

The fluoroscopic barium enema in colonic polyp detection.

The results of 1,500 barium enema studies done by vigorous manual palpation under fluoroscopy were reviewed to determine the adequacy of this technique in detecting polyps above the proctoscopic level. Of the 1,500 patients examined, 103 (7%) were diagnosed radiographically as having a total of 149 polyps. Polyps were found at proctoscopy in about 10% of the 1,500 patients. Colonoscopy or surgery was performed on 35 of the 103 patients. Of the 52 polyps discovered during these procedures, only three were missed at fluoroscopic barium enema study, a false-negative rate of 6%. Fourteen patients whose barium enemas initially were reported as negative for polyps or neoplasia had colonoscopy. Two of the 14 patients had positive findings, for a true-negative rate of 86%. If the detection rate of 7% found at barium enema, taking into account an overlap of 10%, is combined with the rate of 7.5% found at proctoscopy, a frequency of about 14% emerges in detecting polyps in the rectum and in the colon. Thus, the fluoroscopic barium enema when combined with proctoscopy is a sensitive method in the detection of colorectal polyps. In addition, it has the advantage of rapid execution and relatively low cost.

Barium enemas of carcinoma of the colon: sensitivity of double- and single-contrast studies.

A total of 1,140 primary colorectal carcinomas found in 1084 patients during a 5 1/2-year period (1976 to mid-1981) at the Mayo Clinic was reviewed to evaluate the accuracy of detection by the double- and single-contrast barium enema examinations. Both methods were equally sensitive in detecting colon cancer above the proctoscopic level. The error rate (ulcerative colitis excluded) was 4.8% for the single-contrast enema and 4.7% for the double-contrast study. Neither type of examination was superior in finding smaller lesions or earlier staged lesions. Several causes for error were identified: fluoroscopic inexperience, technical factors, misinterpretation of radiologic findings, and distraction.

The role of opiates and endogenous opioid peptides in the regulation of rat TSH secretion.

The administration of morphine to rats at room temperature is reported to suppress serum thyrotropin (TSH) levels by a hypothalamic mechanism. However, it is unknown whether endogenous opioid peptides (EOP) are involved in the control of TSH secretion. The present studies show that naloxone (10 mg/kg, i.p.) an opiate-receptor antagonist, prevented the decline in rat serum TSH which occurs with heat exposure. Morphine sulfate (20 mg/kg, i.p.) treatment prevented the cold-induced elevation in serum TSH and pretreatment with haloperidol (0.3 mg/kg, i.p.) eliminated morphine's influence. Medial-basal hypothalamic thyrotropin-releasing hormone (TRH) content, measured by RIA, increased in the morphine-treated rats which were exposed to 4 degrees C. A submaximal intravenous dose of TRH (300 ng/100 g) was given to determine whether morphine suppresses serum TSH through the release of hypothalamic thyrotropic inhibitors. Morphine pretreatment did not alter TSH stimulation by TRH. Morphine alone or combined with TRH did not alter basal or stimulated TSH secretion in vitro. These studies strongly suggest that, in rats, the EOP modulate TSH secretion under conditions such as acute heat exposure which was associated with a decline in serum TSH. Under specific circumstances, the suppression of serum TSH by morphine may be dopamine-dependent.

Role of growth hormone in experimental phosphorus deprivation in the rat.

The demands of growth are known to exacerbate the effect of phosphorus deprivation (PD). We examined whether changes associated with PD could be prevented in young rats in which growth and growth hormone (GH) were eliminated by hypophysectomy (HPX) and whether PD in normal intact rats (INT) was associated with increased secretion of GH. INT or thyroxine- and ACTH-replaced HPX rats were fed one of the three diets: 0.31% P (NP); 0.027% P (LP), and 0.31% P, pair-fed with LP-mates (NP-PF). The results indicate that HPX did not qualitatively alter several physiologic responses to PD: (a) serum and urinary phosphorus (P) decreased and urinary calcium (Ca) increased; (b) net intestinal Ca retention fell and duodenal sac uptake of 45Ca rose; and (c) external P balance was restored and duodenal sac uptake of 32P-phosphate increased. Only the hypercalcemia seen in INT, LP rats was prevented by HPX. In INT rats serum immunoassayable GH levels, measured in single samples, were not different between different dietary groups while pituitary bioassayable GH was reduced in both LP and NP-PF rats when compared to the NP rats. Thus, except for hypercalcemia, the physiologic responses associated with PD are not prevented by the elimination of growth and GH, and the development of these responses in INT rats was not associated with a consistent or specific alteration in GH secretion.

Effect of human chorionic gonadotropin on thyroid function in euthyroid men.

The effect of large doses of commercial hCG on thyroid function was studied in eight men who received 100,000 or 150,000 IU hCG iv. These large doses of hCG produced definite thyroidal iodine release (TIR) responses in all eight men. The TIR after hCG administration was more delayed and of lesser magnitude than the TIR responses to TSH and TRH. There were no significant changes in serum T4, T3, or TSH for 48 h after hCG administration. No clinical side effects were noted in the subjects after iv administration of these large doses of hCG. The results of this study indicate that hCG is a weak thyroid stimulator in man.

Chronic ulcerative colitis and colon cancer: can radiographic appearance predict survival patterns?

Twenty-four patients with chronic ulcerative colitis and colon carcinoma who underwent barium enema examination prior to the diagnosis of cancer were studied in an attempt to correlate the radiographic appearance of the tumors with their biologic behavior, as well as to assess the accuracy of the barium enema in detection. Of the radiographically detected tumors, 65% displayed an annular infiltrative appearance, with the prime radiographic manifestation being relative nondistensibility of the involved segment. The remaining tumors had various appearances more typical of noncolitic colon cancer. Patients with infiltrative lesions had a much poorer prognosis than those with noninfiltrative lesions. Of 33 gross tumors described pathologically and/or surgically, only 70% were correctly detected by barium enema. It is suggested that periodic barium enema examinations are an unreliable means of following patients with ulcerative colitis if malignant degeneration is to be detected at an early and curative stage. Prophylactic proctocolectomy may be the best therapeutic approach.

Altered responsiveness to hypophysiotrophic hormones of perifused rat pituitary tumours.

Since growth hormone (GH) and prolactin (Prl) secretion by human pituitary tumours is often influenced by the hypophysiotrophic hormones thyrotrophin-releasing hormone (TRH) and somatostatin (SRIF), we have examined the responses of several transplantable rat pituitary tumours to these substances in a perifusion apparatus. The MStT/W15 tumour did not alter its secretion of GH and Prl in response to TRH, SRIF, or a partially purified porcine hypothalamic extract containing GH-releasing activity; normal rat pituitaries show clear responses to each of these substances. Theophylline and dibutyryl cyclic AMP each provoked increased GH and Prl release from the tumour. A second specimen of the MStT/W15 tumour and a specimen of the MStT/W5 tumour behaved in a manner identical to the original MStT/W15, showing no response to TRH or SRIF, but releasing both GH and Prl when theophylline or dibutyryl cyclic AMP was given. The MtT/F4 tumour increased its secretion of GH in response to TRH, 10 mug/ml, and theophylline, but no effect was seen with lower concentrations of TRH or with SRIF; Prl secretion by the F4 tumour was increased by theophylline, but TRH and SRIF had no effect. The autonomy demonstrated in these experimental tumours may be due to a loss of specific hypophysiotrophic hormone receptors or of secretory activating mechanisms.

The hypothalamic-pituitary-thyroid axis.

Although recent investigations have contributed greatly to our understanding of the function and regulation of the hypothalamic-pituitary-thyroid axis, much remains unclear. The influence of suprahypothalamic areas of the brain on hypothalamic function, the nature of thyroid hormone feedback on the hypothalamus, and the physiologis sugnificance of prolactin release by TRH are all topics requiring further study. The extensive experience which has been accululated in the use of TRH as a diagnostic tool has led to its acceptance as a safe, convenient, rapid method of assessment of pituitary and thyroid function. It appears that TRH testing is useful in evaluation of pituitary TSH and prolactin reserve in patients with pituitary lesions; in the differentiation of pituitary and hypothalamic causes of hypothyroidism; in diagnosis of euthyroid Graves' disease; in the evaluation of the adequacy of TSH suppression in thyroid hormone therapy of nodular goiter; and possibly in the diagnosis of mild primary hypothyroidism.