Correlations between clinical presentations of adult trigger digits and histologic aspects of the A1 pulley.

PURPOSE: We aimed to report by light microscopy the normal histology of the A1 pulley, describe the histologic abnormalities of A1 pulleys in trigger digits, and look for possible correlations between these findings and the severity of the disease. METHODS: In a series of 104 trigger digits operated on in 80 adult patients, the A1 pulleys were removed and histologically studied. The findings were compared with 55 normal A1 pulleys obtained from fresh-frozen cadaveric specimens. RESULTS: The normal A1 pulley was composed of 3 layers: layer I, an inner, avascular, concave unicellular or bicellular gliding layer containing cartilage-like cells; layer II, a middle layer, also avascular, characterized by spindle-shaped fibroblasts; and layer III, an outer, richly vascularized layer, continuous with the membranous tendons sheath. We used a 3-grade classification, increasing in severity, to describe the histologic abnormalities observed in trigger digit A1 pulleys. Mild abnormalities (grade 1) were those with a fibrocartilaginous gliding surface almost intact. The margin between the fibrocartilaginous and membranous portions of the pulley was well delineated. In moderate abnormalities (grade 2), the avascular fibrocartilaginous gliding surface appeared fissured and thinner. The inner layer (I) was interrupted and replaced by fibrous tissue, with fissures that did not cross through the middle layer (II). A mild vascular network hyperplasia was observed in the outer layer (III), which began to invade the fibrocartilage. In severe abnormalities (grade 3), the fibrocartilaginous gliding surface was thin, discontinuous, or even completely destroyed. The vascular network hyperplasia became excessive and reached the synovial space of the flexor tendon sheath. The histologic features were correlated with the severity of the clinical symptoms (p < .001). CONCLUSIONS: The histologic abnormalities observed in the A1 pulley of trigger digits are characteristic and not related to inflammation. As the trigger digit worsens, the gliding surface begins to wear and is gradually replaced by a secondary invasive hyperplasia from the outer layer. These abnormalities could be caused by a modification or an increase of the mechanical stresses along the flexor tendons.

Influence of gastrin on human astrocytic tumor cell proliferation.

BACKGROUND: Gastrin and cholecystokinin (CCK) mediate their effects through at least two types of receptors (CCK receptors A and B). While it has been hypothesized that gastrin, a stimulator of gastric acid secretion, is also a neurotransmitter and a stimulator of cell proliferation in various normal and neoplastic tissues, its effect on astrocytic brain tumors has not been actively investigated. PURPOSE: Our goal was to determine the effects of gastrin and gastin and/or CCK antagonists on the proliferation in vitro of astrocytic tumor cells by use of both established cell lines and primary cell cultures of tumor tissue. METHODS: Ten established astrocytic tumor cell lines, SW1088, SW1783, Hs683, H4, U87, U118, U138, U373, T98G, and A172, were studied. The effects of added gastrin (at 0.01, 0.1, and microM) and the gastrin/CCK antagonists L-365,260, CI-988, L-364,718, and JMV 234 (each at 0.01, 0.1, and 1 microM) on the cellular proliferation rates of the 10 cell lines were indirectly measured by use of the colorimetric tetrazolium assay. The influence of gastrin (at 0.01 microM) on the cellular proliferation of primary cultures from nine freshly explanted astrocytic tumors was assessed by means of tritiated thymidine uptake and autoradiography. RESULTS: At specific concentrations, added gastrin increased the cellular proliferation of three established astrocytic cell lines (A172, Hs683, and SW1088), decreased it in two (U373 and T98G), and was without effect on the remaining five. Gastrin decreased cellular proliferation in one primary astrocytic tumor cell culture, stimulated it in five, and had no apparent effect in the remaining three. L-365,260, a CCK receptor B antagonist used at 0.01 microM, increased cellular proliferation in seven cell lines (A172, H4, Hs683, SW1783, T98G, U118, and U138), decreased it in one (U87), and had no effect in the remaining two. CI-988, another CCK receptor B antagonist used at 0.01 microM, inhibited cellular proliferation in five cell lines (A172, H4, SW1783, U373, and U87), stimulated it in two (T98G and U138), and had no effect in three. The CCK receptor A antagonists L-364,718 and JMV 234, both used at 0.01 microM, affected the cellular proliferation of only three of the 10 cell lines. CONCLUSIONS: These results suggest that gastrin (and perhaps CCK that belongs to the same peptide family) may play a role in the growth of a substantial proportion of human astrocytic tumors.

Effect of prolactin and estradiol on cell proliferation in the uterus and the MXT mouse mammary neoplasm.

With the use of an in vivo tritiated thymidine [( 3H]dThd) nuclear labeling followed by autoradiography, the effects at different times before sacrifice of prolactin (PRL) and/or 17 beta-estradiol (E2) were studied in C57BL X DBA/2f)F1 mice given transplants of the MXT hormone-sensitive mammary tumor whose growth was previously shown to be influenced by E2 and/or progesterone. Uteri were chosen as controls for the methodology. Experiments were conducted on ovariectomized mice submitted to endocrine manipulation to achieve plasma PRL modifications. In addition to E2, the proliferation of cancer cells, assessed by the measurement of thymidine labeling indices (TLIs), was demonstrated to be enhanced by ovine prolactin (oPRL) and Sulpiride and strongly slowed down by castration and 2-bromo-alpha-ergokryptin treatment, thus emphasizing the great importance of PRL in mammary cancer development. Moreover, a pulse of 1 mg oPRL/animal produced a marked TLI rise in tumors, lasting from the 6th to the 48th hour after its injection and reaching a maximum at 24 hours. PRL had no proliferative effect on the uterine luminal epithelium. When PRL and E2 were injected concomitantly, the profile of stimulation was quite similar to that obtained with E2 alone; i.e., a maximum stimulation was observed at the 24th and 36th hours after hormonal pulse. From these data it is concluded that, in spayed mice, not only E2 but also PRL is of major importance leading to enhanced proliferation of MXT mammary neoplastic cells. Further investigations are needed to throw light on the cellular events presiding over the action of PRL and E2 at the cancer cell level.

Influence of pituitary grafts or prolactin administrations on the hormone sensitivity of ovarian hormone-independent mouse mammary MXT tumors.

The sensitivity of MXT mouse mammary tumors to ovarian hormones was assessed by the following parameters: growth in vivo; presence or absence of estrogen receptors and progesterone receptors; and histological differentiation. A spontaneous evolution from hormone sensitivity (HS) to hormone independence (HI) was observed when the tumors underwent monthly transplantation onto intact recipient mice, with the tumors fulfilling the criteria of HI tumors after the 12th transplantation. In contrast, the tumors recovered most of the criteria of hormone sensitivity when pituitary isografts were placed under the kidney capsules of HI tumor-bearing animals or when these animals received daily administrations of prolactin over several months. Sensitivity to 17 beta-estradiol, progesterone, or prolactin was further assessed by actinomycin binding on the nucleus and thymidine labeling index, both measured by autoradiography. These technical approaches revealed that 17 beta-estradiol and prolactin stimulated the thymidine labeling index of both HI (despite the lack of detectable estrogen receptors) and HS MXT tumors whereas progesterone influenced only that of HS cancers. The three hormones significantly stimulated [3H]actinomycin D binding within HS tumors but not within HI ones. However, such "HI" tumors were characterized by increased actinomycin binding and thymidine labeling index in comparison with HS neoplasms. Thus, all the data presently reported strongly suggest that prolactin is able to restore the hormone-sensitive phenotype within so-called MXT hormone-independent tumors.

Effects on insulin, prolactin, progesterone, and estradiol on DNA synthesis in organ culture of 7,12-dimethylbenz(a)anthracene-induced rat mammary tumors.

The effect of various hormones or hormone combinations on DNA synthesis was investigated in organ cultures of 20 dimethylbenz(a)anthracene-induced rat mammary tumors. Three tumors were insulin independent and were totally insensitive to all other hormones tested. Seventeen tumors were insulin dependent for DNA synthesis and, in the presence of insulin, displayed variable responses to the other hormones. Nine of 12 such tumors were significantly stimulated by the combination of prolactin and progesterone. Given alone, these hormones were effective in only 25% of the tumors tested. Estradiol used at 2 dose levels, 0.001 or 1.0 mug/ml, acted in a reverse manner to progesterone and proved inhibitory in combination with prolactin in 40% of cases. It was ineffective alone except in 1 of 10 cases in which a stimulatory effect was recorded. A comparison in 4 tumors between estimation of DNA synthesis ([3H]thymidine incorporation into DNA) and colchicine-blocked mitoses demonstrated a good concordance. These results are discussed in terms of variations in the degree of hormone responsiveness of individual tumors and of the known hormone-dependent properties of the 7,12-dimethylbenz(a)-anthracene tumors in vivo.

Estradiol-dependent collagenolytic enzyme activity in long-term organ culture of human breast cancer.

An organ culture method suitable for the maintenance of viable human breast cancer for at least 14 days has been described. This method was applied to a total of 94 breast cancer specimens. It allowed good survival of "soft" tumors of various histological types, with loose connective stroma even in hormone-free medium. In contrast, "scirrhous" cancers showed poor survival in hormone-free medium; viable cells were maintained only at the very periphery of the explants. Supplementation of the medium with insulin (10 mug/ml), ovine prolactin (5 mug/ml), and hydrocortisone (1 mug/ml) in various combinations seemed to induce enlargement of viable cancer cells and moderate loosening of the stroma in some cases. However, it did not improve the survival of central tumor cords in scirrhous explants. Further supplementation of the medium with 17 beta-estradiol (minimum effective dose, 0.1 to 10 ng/ml), although it did not affect soft tumors, markedly improved survival of the cancer cells of scirrhous tumors throughout the whole explants, with evidence of collagen digestion around the neoplastic cells. This was observed in 18 of 20 scirrhous cancers subjected to this treatment. Estradiol need not be present during the whole culture period; the results at 14 days were identical in explants treated with estradiol for the first 7 days only or for the entire period. Addition of purified collagenase during the first 24 or 48 hr of culture resulted in complete dissolution of the collage. After such treatment, culture under the usual conditions resulted in excellent survival of the explants without improvement from hormone supplementation; thus, while estradiol was necessary when collagen was present, it was not longer required after collagen digestion. It can be concluded that breast cancer cells in organ culture are only slightly, or not at all, hormone dependent for survival, provided that they are not restrained by a dense collagen barrier. The estrogen-induced changes allowing survival inside the scirrhous explants strongly suggest the presence of an estrogen-dependent collagenolytic enzyme system in the collagen-rich breast cancers. This system could represent an important component of the hormone dependency of human breast cancer growth.

Selenium deficiency-induced growth retardation is associated with an impaired bone metabolism and osteopenia.

Although the importance of selenium for bone metabolism is unknown, some clinical conditions such as Kashin-Beck osteoarthropathy have been associated with selenium deficiency. Although selenium deficiency induces growth retardation in rats, it has not been established whether this growth inhibition is associated with changes in bone metabolism. We investigated the effect of selenium deficiency on bone metabolism in growing male rats fed a selenium-deficient diet for two generations (Se-). In Se- rats, erythrocyte glutathione peroxidase activity and plasma selenium concentration were strongly reduced compared with pair-fed selenium-adequate rats (Se+). Weight and tail length were reduced by 31% and 13% in the Se- rats, respectively (p < 0.001). The Se- diet was associated with a 68% reduction of pituitary growth hormone (GH; p = 0.01) and a 50% reduction of plasma insulin-like growth factor I (IGF-I; p < 0.001). Plasma calcium was lower and urinary calcium concentration was greater in Se- rats. This group had a 2-fold increase in parathyroid hormone (PTH) and 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] in plasma. Plasma osteocalcin and urinary deoxypyridoline were reduced by 25% and 57% in the Se- rats (p < 0.001). Selenium deficiency resulted in a 23% and 21% reduction in bone mineral density (BMD) of the femur and tibia (p < 0.001) and this effect persisted after adjustment for weight in a linear regression model. A 43% reduction in trabecular bone volume of the femoral metaphysis (p < 0.001) was found in Se- rats. This experimental study shows that growth retardation induced by selenium deficiency is associated with impaired bone metabolism and osteopenia in second-generation selenium-deficient rats.

Identification of high versus lower risk clinical subgroups in a group of adult patients with supratentorial anaplastic astrocytomas.

The present work investigates whether computer-assisted techniques can contribute any significant information to the characterization of astrocytic tumor aggressiveness. Two complementary computer-assisted methods were used. The first method made use of the digital image analysis of Feulgen-stained nuclei, making it possible to compute 15 morphonuclear and 8 nuclear DNA content-related (ploidy level) parameters. The second method enabled the most discriminatory parameters to be determined. This second method is the Decision Tree technique, which forms part of the Supervised Learning Algorithms. These two techniques were applied to a series of 250 supratentorial astrocytic tumors of the adult. This series included 39 low-grade (astrocytomas, AST) and 211 high-grade (47 anaplastic astrocytomas, ANA, and 164 glioblastomas, GBM) astrocytic tumors. The results show that some AST, ANA and GBM did not fit within simple logical rules. These "complex" cases were labeled NC-AST, NC-ANA and NC-GBM because they were "non-classical" (NC) with respect to their cytological features. An analysis of survival data revealed that the patients with NC-GBM had the same survival period as patients with GBM. In sharp contrast, patients with ANA survived significantly longer than patients with NC-ANA. In fact, the patients with ANA had the same survival period as patients who died from AST, while the patients with NC-ANA had a survival period similar to those with GBM. All these data show that the computer-assisted techniques used in this study can actually provide the pathologist with significant information on the characterization of astrocytic tumor aggressiveness.

Characterization of astroglial versus oligodendroglial phenotypes in glioblastomas by means of quantitative morphonuclear variables generated by computer-assisted microscopy.

The current WHO classification places glioblastomas in the astrocytoma category. However, whether or not glioblastomas also show oligodendroglial differentiation remains a matter of controversy. This study investigates, at the morphonuclear level, the hypothesis that some glioblastomas (GBMs) may also represent the ultimate level of malignancy in the oligodendroglial lineage. Using a series of 164 GBMs, we sought to ascertain whether any of these GBMs exhibited phenotypical characteristics that were more closely related to oligodendroglial lineages than astrocytic lineages. Phenotypical features were quantitatively determined by means of the computer-assisted microscope analysis of Feulgen-stained nuclei, a process that made it possible to quantitatively describe the patterns of the cell nuclei (and, more specifically, of their chromatin) through 16 variables, and the distribution of the nuclear DNA content (DNA ploidy) through 8 variables. The phenotypical characteristics typical of astrocytic and oligodendroglial tumors were analyzed by means of Discriminant Analysis, a statistical multivariate analysis, performed on a series of 65 astrocytic and oligodendroglial tumors. This series consisted of 14 WHO grade II and 19 grade III astrocytomas and 24 WHO grade II and 8 grade III oligodendrogliomas. This multivariate analysis enabled an accurate model to be produced that distinguished between astrocytomas and oligodendrogliomas on the basis of 5 cytometry-generated variables. This model was used to characterize the phenotype of each of the 164 glioblastomas. The results show that of these 164 glioblastomas, 6 (about 3.5%) displayed phenotypes that were very similar to oligodendrogliomas, and 141 displayed phenotypes that were very similar to astrocytomas. The phenotypes of the 17 remaining GBMs were too ambiguous to be categorized as having a pure astrocytic or oligodendroglial lineage.

Autonomous secretion of follicle-stimulating hormone by long term organ cultures of rat pituitaries.

Pituitaries removed from ovariectomized adult rats were maintained for 18 weeks in organ culture using three different culture media. Gonadotropin secretion was assessed by RIA and was correlated with the histological features of the cultures. In medium favoring prolonged survival of the cultures, LH content of the medium fell to a low level within a few days. In the same cultures, FSH production initially decreased before increasing and leveling at a plateau which persisted until the end of the culture period. Cultures in medium unsuitable for long term survival of pituitary tissue displayed a similar decrease in LH production along with a gradual fall of FSH. It was concluded that contrary to LH, FSH may be secreted autonomously by pituitaries removed from hypothalamic control, provided that culture conditions are adequate for survival of gonadotropes.

In vitro influence of gastrin, oestradiol and gonadotropin-releasing hormone on HCT-15 and LoVo human colorectal neoplastic cell proliferation.

We set up in vitro several human colorectal neoplastic cell lines that we labelled "hormone-sensitive" (HS) in comparison to the original cell lines which appeared to be rather "hormone-insensitive" (HI). We used LoVo and HCT-15 human colorectal neoplastic cell lines and studied the influence of 17 beta-oestradiol (E2), gastrin and two gonadotropin-releasing hormone (GnRH) analogues, HRF and buserelin, on the proliferation of the HS and HI variants of the LoVo and HCT-15 cell lines. Cell proliferation was evaluated by a colorimetric assay, the MTT test. Our results show that E2, gastrin, HRF and buserelin did not induce a significant stimulatory influence on the HI variants of the LoVo and HCT-15 cells, i.e. the cells that were cultured in a hormone-free 10% FCS-supplemented medium. In sharp contrast, the colorectal cells cultured for 30 passages in an E2 and/or gastrin + 1% FCS-supplemented medium showed a marked tropic response to E2, gastrin, HRF and buserelin. However, the HS variants of the HCT-15 cells appeared less sensitive to the two GnRH analogues than did the HS variants of the LoVo cells.

In vitro characterization of radiotherapy-induced morphonuclear modifications on chemosensitive as opposed to chemoresistant neoplastic cells.

PURPOSE: We describe by means of digital cell image analysis the influence of X-ray radiation on three in vitro cultured cell lines for which we set up chemosensitive and chemoresistant variants. METHODS AND MATERIALS: The three cell lines correspond to the MXT mouse mammary and the T24 and J82 neoplastic human bladder cells. The digital cell image analysis was carried out by computing morphometric (nuclear size), densitometric (proportion of cells in the G2 cell cycle phase), and textural features (chromatin pattern characteristics) on Feulgen-stained nuclei. RESULTS: The results show that such digital cell image analyses make it possible to monitor radiotherapy-induced effects on these morphonuclear characteristics accurately. X-ray radiotherapy induces a dose-dependent increase in the proportion of cells in the G2 phase of the cell cycle along with a decrease in the overall chromatin condensation level. These two concomitant phenomena lead to a marked radiotherapy-induced increase in nuclear size. We also observed that radiotherapy-induced effects at the morphonuclear level are not only highly specific to the cell type analyzed, that is MXT mouse mammary or J82 or T24 human bladder carcinoma cells, but also to the fact that the cells are either chemosensitive or chemoresistant. CONCLUSION: The digital cell image analyses of Feulgen-stained nuclei is helpful in monitoring the irradiation-induced morphonuclear modifications.

Monitoring of radiotherapy-induced morphonuclear modifications in the MXT mouse mammary carcinoma by means of digital cell image analysis.

The present work deals with the characterization of the morphological changes induced by ionizing radiation on MXT mouse mammary cancer cell nuclei. The monitoring of the radiotherapy-induced morphonuclear modifications was carried out by means of digital cell image analysis, which made it possible to compute morphometric (nuclear area), densitometric (nuclear DNA content) and textural (chromatin pattern characteristics) parameters assessed on Feulgen-stained nuclei. The biological material was obtained from serial fine needle-aspirations performed on control tumors and 2Gy and 8Gy irradiated tumors. Digital cell image analyses were assessed at the 7th, 18th, and 28th days-post irradiation. We showed that radiotherapy induced specific morphonuclear modifications that appeared in a dose-dependent and time-dependent manner. The 2Gy and 8Gy radiotherapy doses led to an increase in the mean nuclear area value, this feature being observed at both the 1st and the 4th week post-irradiation. The resultant effect on the chromatin pattern characteristics corresponded to a decrease in overall condensation level as compared to the control cell nuclei. Finally, the mean nuclear DNA content increased at the 1st week post-irradiation, but decreased at the 4th week post-irradiation as compared to the MXT control tumor. Computerized cell image analysis, therefore, appears to be a useful tool in helping to monitor the radiotherapy-induced effects that occur in neoplastic cell nuclei and which can be observed by means of optical microscopy.

Computer-assisted morphonuclear characterization of radiotherapy-induced effects in MXT mouse mammary adenocarcinomas surviving earlier radiotherapy.

PURPOSE: To present the effects of different radiotherapeutic treatments on the morphonuclear characteristics and growth of the MXT mouse mammary adenocarcinoma. METHODS AND MATERIALS: We collected MXT tumor cells by means of fine-needle aspirations during various radiotherapeutic treatments and analyzed the morphological aspects of the cell nuclei by means of the digital cell image analysis of Feulgen-stained nuclei. In addition, we studied the morphonuclear aspects of cells from MXT tumors that had been radioresistant cell enriched. These radioresistant cell-enriched tumors involved MXT tumors that had survived one or two previous radiotherapies. The radiotherapy-induced effects on the morphonuclear characteristics were monitored by means of both monovariate (one-way variance) and multivariate (principal components and step-wise linear discriminant) analyses. RESULTS: The monovariate analyses showed that radiotherapy significantly influenced the values of the parameters relating to nuclear size (nuclear area--NA), the frequency of small dense chromatin clumps (short run length emphasis--SRL) in the nuclei, and the overall chromatin condensation level (local mean--LM). The global effect corresponded to a decrease in the overall chromatin condensation level in the radioresistant cell-enriched MXT tumors. This decrease occurred concomitantly with an increase in the frequency of the small dense chromatin clumps in the nuclei and a decrease in the nuclear area. The multivariate analyses showed that it was possible to quantitate the proportion of "radiosensitive-like" and "radioresistant-like" cell nuclei in the various MXT tumor types under study. CONCLUSIONS: The development of certain morphonuclear parameters, that is, the NA, the SRL, and the LM, could be proposed to predict the response of human tumors to radiotherapy as, indeed, could the quantitation of the proportion of radioresistant cells.

DNA histogram typing in a series of 707 tumors of the central and peripheral nervous system.

The diagnostic value of ploidy level was investigated in a series of 707 tumors (from 707 patients) of the central and peripheral nervous system. The series contained 91 nerve sheath tumors (NSTs), 222 meningiomas (MNGs), 37 primitive neuroectodermal tumors (PNETs), 293 glial tumors of the adult (ATTs), and 64 brain metastases (MTTs). The ploidy level of each tumor was obtained by means of its DNA histogram type (DHT), which was computed (digital cell image analysis) on Feulgen-stained nuclei from archival formalin-fixed paraffin-embedded materials. The data reveal that the measurement of the ploidy level showed a diagnostic value in some of these tumor cases. Indeed, the proportion of highly aneuploid cases significantly (p < 0.05-0.01) decreased according to the sequence PNETs (72%)-->MTTs (64%)-->ATTs (40%)-->NSTs (23%)-->MNGs (9%). Nevertheless, this significant diagnostic value in terms of a distinction between benignity and malignancy only appears when all the tumors are taken into consideration. Indeed, at the level of one individual patient, our study shows that, like traumatic neuromas, schwannomas, or classical meningiomas, completely benign tumors can exhibit a high aneuploidy level. In contrast, some tumors from histopathological groups like brain metastases or PNETs, which are known to be clinically aggressive, could be completely diploid.

Characterization of nuclear DNA content, proliferation index, and nuclear size in a series of 181 meningiomas, including benign primary, recurrent, and malignant tumors.

The characterization of nuclear area, the proliferation index, and nuclear DNA content was carried out by means of digital cell image analysis, which makes it possible to compute morphometric and densitometric features on Feulgen-stained nuclei from archival, that is, formalin-fixed, paraffin-embedded materials. The 181 meningiomas studied included 173 classic (41 meningotheliomatous, 27 fibroblastic, 82 transitional, nine psammomatous, eight angiomatous and six hemangioblastic tumors) and eight malignant meningiomas (three hemangiopericytomas and five tumors that we labeled HFM, that is, tumors exhibiting evidence of histological features of malignancy). The results reveal a strong relationship between incomplete surgical resection and recurrence on the one hand and between the probability of recurrence and histopathological type on the other. Whereas neither nuclear area nor nuclear DNA content assessments were helpful in distinguishing the six classic and the two malignant meningioma subgroups, a statistically significant increase in proliferative activity was observed in the malignant meningiomas as compared with classic ones, excepting hemangioblastomas that proliferate at the same rate as the malignant meningiomas. Furthermore, the multiple meningiomas definitely proliferated more actively than the single ones, but a similar proliferative activity was observed in the nonrecurrent and recurrent meningiomas. Proliferation analyses might be therefore helpful for determining aggressive meningiomas and for planning adjuvant therapy in these cases.

Combined liver-kidney transplantation in primary hyperoxaluria type 1. Bone histopathology and oxalate body content.

In three patients with end-stage renal failure due to primary hyperoxaluria type 1, successful combined liver-kidney transplantation enabled us to assess the insoluble oxalate pool, which was compared with the histopathological changes observed in iliac crest biopsy specimens. Good correlation was observed between the histopathological grade of bone oxalosis and the estimated oxalate content of the body. In the end-stage of oxalate bone disease, hyperparathyroidism does not play a significant role in bone resorption, which appears to be the consequence of the granulomatous reaction induced by oxalate deposition. Combined liver-kidney transplantation should be performed long before this stage. Early hepatorenal grafting in uremia secondary to primary hyperoxaluria type 1 would avoid the deleterious clinical consequences of systemic oxalosis and shorten the duration of postransplant hyperoxaluria, which may compromise the course of kidney graft.

Characterization of factors in routine laboratory protocols that significantly influence the Feulgen reaction.

We investigated the parameters that could affect the cytophotometric analysis of cell nuclei stained by the Feulgen reaction. These parameters included: the hydrolysis temperature (in the normal "room temperature" range); the composition of the Schiff's reagent; the speed of centrifugation of the cell suspensions; the mode of preservation [air-drying or ethanol-formalin-acetic acid (EFA) fixation]; the fixation time; the pronase digestion time; and the concentration of pronase used to obtain cell suspensions from archival (formalin-fixed, paraffin-embedded) materials. Relatively homogeneous material was studied: the MXT mouse mammary adenocarcinoma growing in vivo as tumors with both small and hyperchromatic cell nuclei and in vitro as monolayers with larger and less hyperchromatic cell nuclei. The results of these investigations demonstrate the necessity for the precise definition of a protocol for such procedures as sampling, fixation, and staining of cell nuclei if computerized cell image analyses are to be objective and reproducible. For present purposes this protocol differs depending on whether fresh or archival material is studied. For fresh tissue the protocol is immersion of the sample in EFA within 10 sec, fixation for 30 min, and staining by the Feulgen reaction in which hydrolysis is performed with 6 N HCl at 24 degrees C for 60 min. For archival tissue, the protocol becomes fixation in formol (or EFA), embedding, sectioning at 80 microns, digestion with 0.05% pronase for 2 hr, centrifugation at 1200 x g on glass slides, and staining by the Feulgen reaction in which hydrolysis is performed with 6 N HCl for 60 min at 24 degrees C.

Effects of anterior hypothalamic implantation of gonadal steroids on serum and pituitary gonadotrophins (follicle-stimulating hormone and luteinizing hormone) in the castrated male rat.

Castrated rats were stereotaxically implanted with 1 microng oestradiol benzoate, 5 microng testosterone isobutyrate or, as a control, 10 microng cholesterol, in the hypothalamus. The effects of the steroids on plasma and pituitary gonadotrophins (FSH and LH) were assessed by radioimmunoassay. Our results indicate that, in the male rat, in addition to the arcuate nucleus-median eminence complex, the preoptic suprachiasmatic area is able to control synthesis and secretion of both gonadotrophins, and that it is sensitive to oestradiol and testosterone.

Sensitivity of anterior hypothalamic areas to gonadal steroid implantation in androgenized female rats.

A single injection of 1 mg of a complex of testosterone esters on day 5 of life was used to prepare constantly oestrous rats. Such androgenized female rats were then ovariectomized and submitted to stereotaxical implantation of 1 microgram oestradiol benzoate, 5 microgram testosterone isobutyrate or, as a control, 10 microgram cholesterol in the anterior hypothalamic areas. The effects of the steroids on plasma and pituitary FSH and LH were assessed by radioimmunoassay. As reported previously by us in normal female and male rats, the preoptic-suprachiasmatic area (POA) was able to control synthesis and secretion of both gonadotrophins and did not lose its sensitivity to oestradiol and testosterone in andorgenized rats. Evidence for enhanced prolactin secretion in androgenized rats was derived from immunofluorescence studies of the pituitary gland and from histology of the mammary glands. In this respect the condition of the androgenized females was opposite to that of the males. The present work demonstrated that stimulation of prolactin secretion in androgenized female rats resulted from oestrogen action due to permanent oestrus rather than from impairment of hypothalamo-hypophysial relationships. Indeed, prolactin stimulation was suppressed when the androgenized rats were ovariectomized and restored when they were subsequently implanted with oestradiol in the POA.