Decrease In Proliferative Activity Research Articles

Proliferative Activity of Human Thyroid Cells in Various Age Groups and Its Correlation with the Risk of Thyroid Cancer after Radiation Exposure

The thyroid gland is vulnerable to the carcinogenic effects of ionizing radiation, and there is a well-documented inverse correlation between thyroid cancer and age at exposure, particularly for ages less than 20 yr. One of the factors responsible for this phenomenon may be more rapid cell proliferation in children. The objective of this study was to determine the proliferative rate of normal human thyroid cells in different age groups. We used immunohistochemical analysis to determine the Ki-67 proliferative index in 117 thyroid glands obtained at autopsy, including 25 fetal thyroids (11-40 wk gestation), 55 childhood thyroids (0-19 yr), and 37 adult thyroids (20-60 yr). The rate of Ki-67 labeling in the three groups was 7.4 +/- 6.10, 0.23 +/- 0.15, and 0.08 +/- 0.04% respectively, demonstrating an overall trend for diminishing proliferative activity of thyroid cells with increasing age. However, a lack of correlation was noted between the slopes of cancer risk calculated from previous studies of irradiated populations and proliferative rate in the pediatric age intervals of 0-4 and 5-9 yr, suggesting that other factors are likely to be responsible for the particularly high sensitivity to radiation-induced thyroid cancer among the youngest children. Our findings of a general decrease in proliferative activity of thyroid cells with age may explain, at least in part, the higher risks of radiation-related thyroid cancer in children compared with adults. However, the variation in the rate of cell proliferation is unlikely to be responsible entirely for this phenomenon and other factors may also be involved.

Sirolimus accelerates senescence of endothelial progenitor cells through telomerase inactivation

Background Sirolimus-eluting stent (SES) is commonly used to prevent in-stent restenosis but is not infrequently complicated by late angiographic stent thrombosis (LAST). On the other hand, circulating endothelial progenitor cells (EPCs) play a significant role in the maintenance of endothelial integrity. Aim We examined whether sirolimus affects differentiation, proliferative activity, senescence, colony formation, and network formation in EPCs originated from mononuclear cells (MNCs). Methods and results MNCs were isolated from peripheral blood of healthy volunteers. EPCs outgrew from the culture of MNCs in the presence of vascular endothelial growth factor. When MNCs were incubated with sirolimus at 0.01, 0.1, and 1 ng/ml for 4 days, sirolimus dose-dependently reduced the number of differentiated, adherent EPCs, as assessed by an in vitro culture assay. After ex-vivo cultivation, EPCs became senescent as determined by acidic β-galactosidase staining. When MNCs were treated with sirolimus, sirolimus dose-dependently accelerated the onset of EPCs senescence. RT-PCR analysis demonstrated that FK506-binding protein 12 (FKBP12), a receptor of sirolimus, was expressed in MNCs. To obtain an insight into the underlying downstream effects of sirolimus, we measured telomerase activity and the expression of p27 kip1. Sirolimus decreased telomerase activity dose-dependently, which was accompanied with down-regulation of the catalytic subunit, telomerase reverse transcriptase (TERT). Furthermore, sirolimus up-regulated the cell cycle inhibitor p27 kip1. Having demonstrated that sirolimus accelerated the onset of senescence, we examined whether that translated into a decrease in proliferative activity and clonal expansion. Both MTS assay and BrdU incorporation assay have shown that sirolimus treatment significantly diminished the proliferative activity in EPCs. In addition, colony forming unit assay revealed that sirolimus dramatically decreased colony formation as compared to control (no treatment). Finally, in a Matrigel assay, EPCs treated with sirolimus were shown to be less integrated into the network formation than control (no treatment). Conclusion The inhibitory effects of sirolimus on circulating EPCs potently may affect re-endotheliazation after SES implantation.

Modulation of Hypoxia in Murine Liver Metastases of Colon Carcinoma by Nicotinamide and Carbogen

There is increasing evidence that modulation of tumor hypoxia may improve therapy outcome. However, most preclinical data are derived from subcutaneous rather than orthotopic tumor models. We investigated the effect of the hypoxia-modulating agents nicotinamide and carbogen on tumor hypoxia, tumor blood perfusion, and proliferative activity in liver metastases of the murine colon carcinoma line C26a. In untreated C26a liver metastases, we observed a considerable amount of hypoxia, similar to the amount in liver metastases of patients with colorectal cancer. Compared to untreated mice, we observed a significantly smaller hypoxic fraction in the liver metastases of mice treated with nicotinamide and carbogen breathing as single treatments or in combination. In the group of mice that underwent carbogen breathing, perfusion was significantly lower than in the untreated group, but the decrease was only marginal. The proliferative activity was similar in all groups. In C26a subcutaneous tumors, a similar effect on hypoxia has been observed that was, however, combined with a decrease in proliferative activity. The different effects of nicotinamide and carbogen on parameters of the tumor microenvironment in liver metastases and subcutaneous tumors suggest that the host tissue influences the mechanism by which nicotinamide and carbogen exert their effects. Since tumor hypoxia may be a clinical problem in colorectal liver metastases, our results open possibilities for further research on the effect of hypoxia modifiers on colorectal liver metastases to improve therapy outcome.

Investigations of Interactions of Chlormezanone Racemate and Its Enantiomers on Human Keratinocytes and Human Leucoytes in vitro

Chlormezanone is a centrally acting muscle relaxant introduced in human therapy as a racemic substance. The following investigation was performed in order to investigate whether the racemate and both enantiomers differ in their potential cytotoxicty in vitro. We investigated antiproliferative effects and cytotoxicity (PicoGreen and ATP assay) for human HaCaT keratinocytes, production of oxygen radicals (ROS) by human interleukin-3-stimulated leukocytes (Lucigenin assay) and production of sulfoleukotrienes (Cellular Antigen Stimulation Test – CAST) by human leukocytes. In the dosage range of 0.001 to 0.1 mg/ ml chlormezanone, no antiproliferative effects were measured with the racemate and both enantiomers. At 1.0 mg/ml, a decrease of proliferative activity was seen after 48 h incubation time of about 50% for the enantiomers and of about 80% for the racemate (PicoGreen) and 50% (enantiomers) or 21% (racemate) in the ATP assay, respectively. ROS production was significantly inhibited at concentrations ≤0.01 mg/ ml by the racemate and the (+)-enantiomer, whereas the (–)-enantiomer was less effective. There was no stimulation of sulfidoleukotrienes in human leukocytes by chlormezanone. Present data argue for absence of significant cytotoxicity against human HaCaT keratinocytes and a dose-dependent suppression of ROS production by human leukocytes that is not uniform among the racemate and its enantiomers.

Association of increased estrogen receptor β2 expression with parity-induced alterations in the rat mammary gland

In this study, we investigated the cellular and molecular events involved in parity-related alterations in mammary gland (MG) proliferation and differentiation. Rat MGs were removed on day 9 of either first (nulliparous), second (primiparous) or third (multiparous) pregnancy. Expression of steroid hormone receptors along with cellular biomarkers of proliferation and differentiation were quantified in all MG tissue compartments by immunohistochemistry. Wnt-4 (a Wingless-like morphogenic gene involved in MG development), ERβ and ERβ2 mRNA were evaluated by RT-PCR analysis. Serum levels of mammotrophic hormones were measured. In comparison to nulliparous and primiparous rats, multiparous animals exhibited decreased luminal cell proliferation and PR levels, whereas α-lactalbumin, ERα, ERβ and ERβ2 expression were increased. In myoepithelial cells, while parity induced a decrease in proliferative activity, subsequent pregnancies and lactations lead to an increased state of differentiation. Our results showed that at least two periods of pregnancy and lactation were necessary to modify the studied parameters. The lower proliferative activity and higher differentiation state of the multiparous MG are associated with both a decreased PR expression and increased ERα and ERβ expression. Since ERβ and/or ERβ2 isoform expression was related to parity history, results suggest that the decreased proliferative activity and PR expression observed in the MG of multiparous animals may be associated with overexpression of ERβ and/or the ERβ2 isoform, thereby antagonizing the proliferative effects associated with ERα.

Hormone-induced protection against breast cancer.

Reproductive history is a consistent risk factor for human breast cancer. Epidemiological studies have repeatedly demonstrated that early age of first full-term pregnancy is a strong protective factor against breast cancer and provides a physiologically operative model to achieve a practical mode of prevention. In rodents, the effects of full-term pregnancy can be mimicked by exposure to low doses of estrogen and progesterone or treatment with human chorionic gonadotropin. The cellular and molecular mechanisms that underlie hormone-induced refractoriness are largely unresolved. Several hypotheses have been proposed to explain the protective effects of hormones. These involve the induction of differentiation of the mammary gland to provide a less responsive cell population to carcinogens, a decrease in proliferative activity in the parous gland compared to the age-matched virgin, an altered hormonal environment mediated by a decrease in circulating growth hormone, and an alteration in cell fate mediated by specific molecular changes induced by estrogen and progesterone. The evidence for and against these hypotheses is discussed along with recent results on possible molecular alterations that may underlie the refractory state. One central question that is still unresolved is whether the refractoriness is intrinsic to the mammary epithelial cells and/or mediated by persistent alterations in the host environment.

Proliferation and apoptosis in the epithelium of the developing human cornea and conjunctiva

To determine the distribution of proliferating and apoptotic cells in the human cornea during prenatal and early postnatal development, we examined sections of the bulbar conjunctiva, the limbus as well as the central and peripheral cornea between 11 weeks of gestation and 6 months after birth. The objective was to localize dividing cells by proliferating cell nuclear antigen-like immunoreactivity (PCNA-LI) and apoptotic cells by terminal transferase-mediated nick-end labeling (TUNEL). Before the 17 th gestational week, PCNA-LI was absent in all 4 regions examined, indicating negligible cell proliferation during early development. After 20 weeks, strong PCNA-labeling was observed in all regions examined suggestive of high proliferative activity not only in the limbus and the bulbar conjunctiva, but also in the central and peripheral cornea. This rise in proliferative activity was followed by a steady decline: after 28 weeks, anti-PCNA staining gradually disappeared in the central and peripheral cornea, so that, at 6 months after birth, it was confined to the limbus and the bulbar conjunctiva, resembling the picture described for the adult cornea. TUNEL-positive cells were virtually absent in all 4 regions examined before the 38 th gestational week. Apoptotic cells only started to appear at 38 weeks; at this stage, they were confined to the bulbar conjunctival epithelium. At 6 months after birth, TUNEL-positive cells were observed in the bulbar conjunctival epithelium and the entire cornea; the limbus, however remained devoid of apoptotic cells throughout the entire prenatal and early postnatal period. The present study for the first time localizes proliferating and apoptotic cells in the epithelium of the developing human cornea. Three stages of development can be distinguished: Minimal proliferation (until 17 th week), vigorous proliferation over the entire cornea including the limbus and the bulbar conjunctiva (until 28 th week) and gradual decrease in proliferative activity (after 28 th week) accompanied by the appearance of apoptotic cells.

The chondrogenic potential of periosteum decreases with age

Periosteum contains undifferentiated mesenchymal stem cells that possess the potential for chondrogenesis during cartilage repair and in fracture healing. With aging, the chondrogenic potential of periosteum declines significantly. An organ-culture model was used to investigate the relationship between the chondrogenic potential of periosteum and aging. A total of 736 periosteal explants from the proximal medial tibiae of 82 rabbits, aged 2 weeks to 2 years, were cultured in agarose suspension conditions conductive for chondrogenesis, and analyzed using histomorphometry, collagen typing, wet weight measurement, 3H-thymidine and 35S-sulfate uptake, autoradiography, and PCNA immunostaining. The rabbits were skeletally mature by 6 months and stopped increasing in weight by 12 months. Chondrogenesis declined significantly with age ( P<0.0001) and was maximal in the 1.5–2 month-old rabbits. Explants from the 6 month-old rabbits formed 50% less cartilage, and by 12 months chondrogenesis reached a steady state minimal level. In parallel with this decrease in chondrogenic potential similar decreases were measured in 3H-thymidine uptake ( P<0.0001), 35S-sulfate uptake ( P=0.0117), as well as the thickness ( P<0.0001) and the total number of cells in the cambium layer of the periosteum ( P<0.0001). Autoradiography with 3H-thymidine and PCNA immunostaining confirmed the measured decrease in proliferative activity in the cambium layer where the chondrocyte precursors reside, although the percentage of proliferating cells did not change significantly with age. The most dramatic change was the marked decrease (87%) in the thickness and total cell number in the cambium layer of the perisoteum between the 2 and 12 month-old rabbits ( P<0.05). These data confirm a decline in the chondrogenic potential of periosteum with aging. Thus, one possibility for improving cartilage formation by periosteal transplantation after skeletal maturity would be to stimulate an increase in the total number of cells in the chondrocyte precursor pool early during chondrogenesis.

Structure-bioactivity of C-terminal pentapeptide of osteogenic growth peptide [OGP(10-14)

The amino acid sequence of osteogenic growth peptide (OGP) consists of 14 residues identical to the C-terminal tail of histone H4. Native and synthetic OGP are mitogenic to osteoblastic and fibroblastic cells and enhance osteogenesis and hematopoiesis in vivo. The C-terminal truncated pentapeptide of OGP, H-Tyr-Gly-Phe-Gly-Gly-OH [OGP(10-14)], is a naturally occurring osteoblastic mitogen, equipotent to OGP. The present study assesses the role of individual amino acid residues and side chains in the OGP(10-14) mitogenic activity which showed a very high correlation between osteoblastic and fibroblastic cell cultures. Truncation of either Tyr10 or its replacement by Ala or D-Ala resulted in substantial, but not complete, loss of activity. Nevertheless, only a small loss of activity was observed following removal of the Tyr10 amino group. No further loss occurred consequent to the monoiodination of desaminoTyr10 on meta-position. However, a marked decrease in proliferative activity followed removal of the Tyr10 phenolic or the Phe12 aromatic group. Loss of activity of a similar magnitude also occurred subsequent to replacing Gly11 with L- or D-Ala. Approximately 50% loss of mitogenic activity occurred subsequent to truncation of Gly14 or blocking the C-terminal group as the methyl ester. All other modifications of the C-terminus and L- or D-Ala substitution of Gly13 resulted in 70-97% decrease in activity. Collectively, these data suggest that the integrity of the pharmacophores presented by Tyr and Phe side chains, as well as the Gly residues at the C-terminus, are important for optimal bioactivity of OGP(10-14).

Proliferative activity of early hemopoietic precursors under conditions of increased and decreased activity of protein kinase C in mouse bone marrow

Activation of protein kinase C (transfer from cytosol to cell membranes) with phorbol ester increases proliferation of splenic colony-forming units in the bone marrow. Subsequent incubation of bone marrow cells with the factor inhibiting proliferative activity of early hemopoietic precursors results in the recovery of protein kinase C activity in cell cytosol and a decrease in proliferative activity.

Neuroblastoma cells can actively eliminate supernumerary MYCN gene copies by micronucleus formation--sign of tumour cell revertance?

Human neuroblastoma cell lines frequently exhibit MYCN amplification and many are characterised by the presence of morphologically distinct cell types. The neuronal cells (N-cells) and the so-called flat cells (F-cells) are thought to represent manifestations of different neural crest cell lineages and are considered to be the consequence of neuroblastoma cell pluripotency. In this study, various neuroblastoma cell lines were examined for micronuclei. In F-cells of neuroblastoma cell lines with extrachromosomally amplified MYCN, we observed the frequent occurrence of micronuclei. Using fluorescence in situ hybridisation (FISH) with a MYCN specific probe, we demonstrated that these micronuclei were packed with MYCN hybridisation signals. In addition, in a minor percentage of cells, MYCN signals occurred in clusters, adhered to the nuclear membrane and aggregated in nuclear protrusions. In F-cells, a substantial reduction or lack of amplified MYCN copies was observed. These observations let us conclude that extrachromosomally amplified genes can be actively eliminated from the nucleus resulting in a dramatic loss of amplified sequences in the F-cells. Moreover, reduction or loss of amplified sequences in F-cells was shown to be accompanied by downregulation of MYCN expression, by a decrease in proliferative activity and by upregulation of molecules of the major histocompatibility complex class I (MHC I). Interestingly, F-cells are not restricted to neuroblastoma cell cultures, but also occur in cell lines of other tissue origin. All F-cells share important biological features, interpreted as cell revertance, i.e. loss of the malignant phenotype and properties. This fact, together with the demonstration that neuroblastoma cells do not differentiate into Schwann cells in vivo [1] Ambros et al. NEJM 1996, 334, 1505-1511, do not support the hypothesis that F-cells represent Schwannian/glial differentiation in vitro. We therefore postulate that the elimination of amplified MYCN gene copies in cultivated neuroblastoma cells is in line with the phenomenon of tumour cell revertance.

Cell proliferation and apoptosis during prostatic tumor xenograft involution and regrowth after castration

The biological mechanisms involved in androgen-dependent and -independent prostate cancer growth after castration were analyzed in the LuCaP 23.1 human prostate cancer xenograft model. Athymic mice (n = 82) bearing LuCaP 23.1 xenograft were castrated and tumors were harvested at different time points from day 0 to day 112 post castration. In each group of mice, tumor growth rate (TGR), serum PSA concentration, percentage of tumor cells incorporating bromodeoxyuridine (BUdR index), percentage of apoptotic tumor cells assessed by morphological analysis (apoptotic index), and presence of apoptosis-related DNA "ladder" were analyzed. Castration induced a significant decrease in TGR and serum PSA from day 1 to day 7, and a progressive increase in the 2 parameters from day 14 to day 112, heralding androgen-independent tumor relapse. Meanwhile the BUdR and apoptotic indexes varied as follows after castration: an increase was noted for both at day 3, a significant increase in apoptotic index with a decrease in BUdR index from day 5 to day 14, and a progressive decrease in apoptotic index while BUdR index remained at 50% of the pre-castration value from day 28 to day 112. DNA ladder was present sparsely in tumors grown in non-castrated hosts, universally present in tumors from day 1 to day 28 post castration, and frequent in tumors from day 56 to 112. Castration-induced effects in LuCaP 23.1 tumors were characterized by an increase in number of apoptotic cells and a decrease in proliferative activity. The androgen-independent tumor relapse after castration was associated with a low apoptotic index with no increase in proliferative activity.

Early decrease in proliferative activity of hepatocytes after tips placement: Evidence for tips-induced liver damage: [formula omitted], H. Louis, C. Degraef, O. Le Moine, J. Devière, P. Galand. Dept. of Gastroenterology & Lab. Cyt. Cancer Exp., Erasme Hospital, Brussels, Belgium

Early decrease in proliferative activity of hepatocytes after tips placement: Evidence for tips-induced liver damage: [formula omitted], H. Louis, C. Degraef, O. Le Moine, J. Devière, P. Galand. Dept. of Gastroenterology & Lab. Cyt. Cancer Exp., Erasme Hospital, Brussels, Belgium

Physeal distraction and cell proliferation in the growth plate

We studied the cellular response to physeal distraction in the growth plates of skeletally immature rabbits. We used a new method of labelling and detection of proliferating cells with bromodeoxyuridine (BUdR) and an anti-BUdR antibody. The application of an external fixator but no distraction force produced no changes in the growth plates. After five days of distraction at a maximum force of 20 N, the growth plate became thicker, mainly because of an increase in the number of hypertrophic chondrocytes, but there was no evidence of increased cell proliferation. Recent fractures were seen at the junction of growth plate and metaphysis but the increase in bone length was insignificant. After ten days of distraction at the same maximum force, the chondrocyte columns had become disorganised and cell proliferation was significantly decreased. There was an increase in bone length due to distraction of the fracture gap. In this model, physeal distraction did not stimulate cell proliferation, but actually inhibited it. The apparent increase in growth-plate thickness produced by distraction is not due to increased cell production, but results from inhibition of endochondral ossification and the consequent accumulation of hypertrophic chondrocytes. Any growth after distraction depends on the ability of growth-plate chondrocytes to divide. The decrease in proliferative activity which we found after ten days of distraction suggests the need for caution in the use of such procedures in young children.

Time Course of 2,3,7,8-Tetrachlorodibenzo- p-dioxin (TCDD)-Induced Thymic Atrophy in the Wistar Rat

Exposure to sublethal doses of 2,3,7,8-tetrachlorodibenzo- p-dioxin (TCDD) in Wistar rats results in thymic atrophy and reduced thymic cellularity. In a first experiment, rats were once orally intubated with 0, 1, 5, 25, 50, and 150 μg TCDD/kg body wt and sacrificed at Day 10. The dose that produced one-half of the maximal thymic involution was estimated at about 10-20 μg/kg. The time course (Days 0-21) of thymic atrophy induced by a single oral intubation of 25 μg TCDD/kg body wt revealed four phases. In phase 1 (initiation phase, Days 0-2) a decrease in proliferative activity was seen in cortical thymocytes, whereas no changes occurred in thymic cellularity. Phase 2 (lymphodepletion phase, Days 2-8) was characterized by an initial strong depletion of immature CD4 +CD8 + double-positive (DP) cells (Day 4), followed by a more gradual decrease in the number of mature thymocytes (Day 6). On Day 8 the lymphodepletion was maximal. In phase 3 (stationary phase, Days 8-13) no changes occurred in thymic cellularity. Although the first signs of recovery were already seen on Day 6, indicated by a recovery in proliferative activity in the thymus cortex, an increase in thymic cellularity was observed first after Day 13 (phase 4, recovery phase, Day 13 onward). Reversibility of thymic atrophy is therefore observed within the estimated half-life of TCDD in the rat thymus (>16 days). We conclude that TCDD exerts a rapidly reversible effect on an intrathymic cortical target cell population.

Cell phenotypes and differentiative transitions in mouse submandibular salivary gland defined with monoclonal antibodies to mammary epithelial cells.

Recognition of distinct cell phenotypes within a given organ is important in defining cell relationships during development and in analyzing the role of cell-cell and cell-matrix interactions in growth and differentiation. Phenotypic definition of dissociated heterogeneous cell populations is also essential for studies on mechanisms regulating expression of cell lineage-specific gene products. Mouse submandibular salivary gland (SSG) cell phenotypes in the course of differentiative transitions in vivo and after enzymatic dissociation in primary culture were defined with monoclonal antibodies (MAb) to mammary epithelial cells and polyclonal antibodies to functional cell products. Proacinar cells and differentiating and mature acinar cells were uniquely recognized by an MAb designated 50B8. Ductal cell components were uniquely recognized by an MAb designated JSE3. JSE3 immunoreactivity was particularly useful for detecting the emergence of two SSG duct cell phenotypes, striated ducts and the hormone-responsive granular convoluted tubules (GCTs). JSE3-positive striated duct-like cells were visualized as early as Day 2 after birth and emergence of GCT-like structures from striated ducts was apparent between Days 10 and 11. Differential reactivity of acinar and ductal cells in the developing SSG with either MAb 50B8 or JSE3 suggests the existence of intermediate progenitor cells restricted in their differentiation potential. An interesting pattern of immunoreactivity was observed with an MAb designated 33A10. During the first 2 weeks of SSG postnatal development, shared 33A10 immunoreactivity was observed among proacinar and differentiating acinar cells and all differentiating ductal segments. Coincident with a decrease in proliferative activity at about Day 18, 33A10 immunoreactivity became restricted to the GCT cell lineage before the appearance of GCT functional products, epidermal and nerve growth factors. Although the SSG antigen recognized by MAb 33A10 is presently undefined, its expression pattern suggest a molecule with a dual role in development and growth events and in hormone-dependent secretory function. Advantage was taken of the observed differential immunoreactivities to define the phenotypic identity of dissociated, mature SSG cells before and after culture. Dissociated SSG fractions enriched for either JSE3- or 50B8-positive cells could be maintained in short-term cultures without loss of expression of duct- or acinar cell-specific immunoreactivity. In addition to providing markers for defining dissociated SSG cells before and after culture, the described immunoreactivities may permit separation or enrichment of an early duct cell population before its commitment to a specific cell lineage. This approach may also provide the means to define regulatory signals involved in the differentiation of intermediate progenitor cells.

Flow cytometric analysis of in vitro micronucleus induction in hepatocytes treated with carcinogens

The micronucleus test is frequently used in established cell lines to detect genotoxic chemicals. In contrast, there is only very limited experience concerning the applicability of this test in primary cultures of hepatocytes. The induction of micronuclei (MN) by methyl methanesulphonate (2 m m) and by the indirect carcinogens cyclophosphamide (CP, 0.4–4 m m) and diethylnitrosamine (DEN, 1–10 m m) has therefore been studied in rat liver cells in vitro. Analysis and quantification of MN, as well as determination of the proliferative activity of the hepatocytes, was performed by flow cytometric techniques. All three chemicals increased the frequency of MN at incubation times of more than 48 hr. The relative increase in MN compared with that in untreated or solvent-treated cultures, however, was at the most only three-fold, since the frequency of MN increased markedly in the control cultures also. There was a marked decrease in proliferative activity of the hepatocytes, as shown by the decrease in frequency of cells in the second G1-phase at the highest concentration of CP and at all concentrations of DEN. In conclusion, flow cytometric analysis of MN enables a fast and reliable determination of cytogenetic effects in hepatocyte cultures treated with chemicals. However, the large number of MN in untreated hepatocytes, which is possibly a consequence of DNA damage induced by the isolation procedure, may limit the sensitivity of the method.

Interleukin-1 inhibits human thyroid carcinoma cell growth.

In order to further clarify the growth regulating effect of cytokines on thyroid cell proliferation, the effect of interleukin-1 (IL-1) was tested on growth of human papillary and follicular thyroid carcinoma cells (NPA and WRO). Although fetal bovine serum stimulated DNA synthesis, TSH did not affect cell growth and the level of thyroglobulin messenger RNA (mRNA) in NPA. Eight-bromo-cAMP and forskolin, however, significantly suppressed DNA synthesis, suggesting the impairment of TSH receptor signal transduction in NPA cells despite of the presence of [125I]TSH binding. Both IL-1 alpha and beta inhibited [3H] thymidine uptake into NPA cell DNA in a dose- and time-dependent manner at concentrations ranging from 5 x 10(2) to 10(5) U/L; 10(5) U/L of IL-1 suppressed DNA synthesis by more than 40% of control. The suppressive activity of IL-1 beta was more potent than that of IL-1 alpha in spite of the same lymphocytes activating factor (LAF) activity. Steady state levels of c-myc mRNA transcripts were also inhibited by both IL-1 alpha and beta, respectively. On the other hand, IL-1 alpha and beta did not affect the WRO cell growth. Although forskolin stimulated cAMP production in NPA cells, IL-1 did not induce cAMP generation. Indomethacin (1.0 approximately 10 mg/L) did not alter the suppressive activity of IL-1. Neither IL-1 alpha and beta caused a cytotoxic effect on the NPA cells. Although changes in c-myc mRNA content may just be an epiphenomenon of the decrease in proliferative activity in response to IL-1, and not a direct target of action of the peptide, these results demonstrate the inhibitory effect of IL-1 on cell growth of NPA. IL-1 may have an aspect of antitumorigenic action in some human thyroid carcinoma differentiation and replication.

Serosal patching impairs intestinal adaptation following enterectomy

Increasing intestinal absorptive surface by mucosal regeneration on serosal patched intestinal defects is a potential surgical treatment for the short bowel syndrome. We previously found in long-term studies that serosal patching in dogs undergoing 75% enterectomy was deleterious to intestinal adaptation and absorption. Our aim was to evaluate the effect of serosal patching on the early morphologic and functional changes in postresectional adaptation and to examine the role of polyamine metabolic pathways in this process. Five unoperated New Zealand white rabbits (GP I) served as controls. Twelve other rabbits underwent either 50% distal enterectomy alone (GP II) or simultaneously had two 2 × 5-cm full-thickness ileal defects patched with adjacent cecal serosa (GP III). Animals in GP II gained an average of 7.2 ± 5.3% body weight, whereas GP III animals lost 5.6 ± 9.0% body weight ( P < 0.05). Intestinal remnant length was significantly less in GP III 3 weeks postoperatively (66 ± 11 vs 85 ± 8 cm, P < 0.05) as was mucosal protein content (4.1 ± 1.8% vs 6.2 ± 1.9%) but villus height was similar in GPs II and III (505 ± 131 vs 508 ± 110 μm). In vitro mucosal function was similar in all three groups. Crypt cell production rate was significantly lower while ornithine decarboxylase and diamine oxidase activity were higher in GP III compared to GP II. However, polyamine levels were similar in all three groups. Serosal patching impairs intestinal adaptation following massive enterectomy. This is due in part to a decrease in proliferative activity which is not directly related to altered polyamine levels. Thus, procedures intended to improve intestinal function after massive intestinal resection should not be performed until intestinal adaptation is complete.

Proteoglycans synthesized by gingival fibroblasts derived from human donors of different ages.

The proteoglycans synthesized by fibroblasts derived from human donors of ages ranging from 12 years to 68 years have been studied. In addition, the in vitro proliferation rates of the various cell strains were studied and demonstrated that increasing donor age correlated with a decrease in proliferative activity. The incorporation of [35S]-sulfate into proteoglycans decreased with increasing donor age with cells from the oldest donor demonstrating a 50% reduction compared with cells from the youngest donor. Analysis on Sepharose CL-4B of isolated [35S]-labeled proteoglycans for molecular size distribution revealed few differences between the cell-layer-associated proteoglycans of all cell strains studied. However, analysis of the medium-associated [35S]-labeled proteoglycans demonstrated an increase in the amount of small molecular size proteoglycans with increasing age. More specific analysis of the glycosaminoglycan composition revealed an increase in heparan sulfate from 52% to 73% in the cell-layer-associated proteoglycans of cells from the youngest and oldest donors, respectively. Accompanying this increase was a relative decrease in dermatan and chondroitin sulfate content from 24% to 13% and 25% to 16%, respectively, with increasing donor age. Additionally, the degree of N-sulfation of cell layer heparan sulfate increased with age. Heparan sulfate levels increased in the medium as well with increasing age, with a concomitant decrease in chondroitin sulfate. The quantity of medium-derived dermatan sulfate remained relatively evenly distributed throughout the various ages studied. The various differences noted are considered to reflect the general metabolic changes associated with aging. In particular the increase in heparan sulfate content with age is considered to be related to the decreased proliferative activity of the fibroblasts with increasing age.