Effects of pre- and postnatal cysteamine exposure on renal function in the rat.

The safety of cysteamine after renal transplantation and during pregnancy is an important issue, since girls with cystinosis are in better health on cysteamine therapy and thus more likely to become pregnant. In the first study, cysteamine was given to pregnant rats on days 6.5-18.5 post conception in oral doses of 0, 37.5, 75, 100, and 150 mg/kg per day. The dams were sacrificed on day 20.5, the fetal kidneys removed and prepared for histological examination. In the second study, cysteamine was given to dams on days 6.5-19.5 post conception in oral doses of 0, 37.5, 50, and 75 mg/kg per day. Dams were allowed to give birth naturally and pups were given cysteamine on days 4-21 to yield the same oral doses of cysteamine given to the dam. Renal function was evaluated on day 35. Histological examination of fetal kidneys revealed no changes even in kidneys from fetuses with growth retardation and malformations. Furthermore, there were no alterations in renal function in offspring on day 35. These findings demonstrate that cysteamine therapy does not affect renal development in the rat. Further investigations will be required to prove whether cysteamine therapy has the potential to affect renal development in the human.

Expression of cysteine proteases in extraembryonic tissues during mouse embryogenesis.

The expression of cathepsin B- and L-specific mRNAs as well as active forms of the enzymes was determined in mouse placenta and visceral yolk sac from 7.5 through 17.5 days postconception, a period marked by major anatomic transitions in the mouse conceptus. The level of specific mRNA was determined relative to the 28S ribosomal RNA in a series of multiprobe ribonuclease protection assays using high-specific-activity antisense cathepsin B and L riboprobes. The molecular forms of active cysteine proteases present in the tissues at the time of extraction were detected using a membrane-permeant radiolabeled active site-specific inhibitor, Fmoc-[(125)I(2)]Tyr-Ala-CHN(2). The results of this study show that the expression of active cathepsin L relative to active cathepsin B is significantly higher in visceral yolk sac than in placenta, consistent with a higher proteolytic requirement for the former tissue. Active cathepsin L was highest at Day 9.5 in visceral yolk sac, a stage at which it has been shown that proteolysis in this organ is required for production of amino acids for embryonic protein synthesis. Cathepsin L mRNA was also elevated in the Day 9.5 placenta, but paradoxically this did not result in an increase in cellular active enzyme. An unknown protein, termed p14, highly expressed in placenta, also reacted with the inhibitor. Expression of this protein was highest early during gestation in the ectoplacental cone, suggesting that p14 may be important in the implantation process.

Psychiatric features of 36 men convicted of sexual offenses.

BACKGROUND: To increase understanding of the relationship between sexual violence and mental illness, the authors assessed the legal histories and psychiatric features of 36 males convicted of sexual offenses. METHOD: Thirty-six consecutive male sex offenders admitted from prison, jail, or probation to a residential treatment facility received structured clinical interviews for DSM-IV Axis I and II disorders. The participants' legal histories, histories of sexual and physical abuse, and family histories of psychiatric disorders were also assessed. RESULTS: The participants' mean +/- SD age was 33+/-8 years. They had been convicted a mean of 1.8+/-1.4 times (range, 1-9 times) for sexual offenses and incarcerated a mean of 8+/-6 years (range, 0-22 years). Participants displayed high rates of lifetime DSM-IV Axis I disorders: 30 (83%) had a substance use disorder; 21 (58%), a paraphilia; 22 (61%), a mood disorder (13 [36%] with a bipolar disorder); 14 (39%), an impulse control disorder; 13 (36%), an anxiety disorder; and 6 (17%), an eating disorder. Participants also displayed high rates of Axis II disorders, with 26 (72%) meeting DSM-IV criteria for antisocial personality disorder. In addition, subjects reported experiencing high rates of sexual (but not physical) abuse and high rates of Axis I disorders, especially substance use and mood disorders, in their first-degree relatives. Compared with subjects without paraphilias, subjects with paraphilias displayed statistically significantly higher rates of mood, anxiety, and eating disorders, as well as significantly higher rates of childhood sexual abuse. CONCLUSION: Recognition and treatment of major psychiatric disorders among sex offenders may increase chances for successful rehabilitation, reduce recidivism and public victimization, and produce significant public health and economic benefits. More studies in this area appear warranted to search for more effective interventions for this severe public health problem.

Evaluation of the reproductive and developmental safety of cysteamine in the rat: effects on female reproduction and early embryonic development.

Cystinosis is an autosomal recessive metabolic disease in which the amino acid cystine accumulates in lysosomes due to a defect in lysosomal cystine transport. Cystinosis in infancy is associated with poor growth, muscle wastage, and death at about age 10 due to kidney failure. Treatment with cysteamine and kidney transplantation enables cystinotic girls to reach reproductive age and to be healthy enough to permit pregnancy. It is not known whether exposure to cysteamine will have adverse effects on reproduction in the human. It is also possible that some of the complications seen in cystinotic children could be avoided if a pregnant woman carrying a cystinotic fetus were given cysteamine. However, this treatment is not likely to occur until therapeutic exposures to cysteamine are judged to present no increased risk to the human fetus. As part of a larger investigation assessing the reproductive and developmental safety of cysteamine (as phosphocysteamine) using the rat, the two studies reported herein were performed. The first, a dose-finding study, led to the selection of 150 mg/kg/day as the highest dose of cysteamine used for the second and primary focus of this report. The second study involved the exposure of female rats to cysteamine from premating through day 6.5 postconception and assessment of female fertility and early embryonic development. Cysteamine was administered orally in doses of 0, 37.5, 75, 100, or 150 mg/kg/day. There were no clinical signs of maternal toxicity during the exposures of 2 to 5 weeks before successful mating. Animals in the 150 mg/kg/day group experienced a nonsignificant decrease in body weight gain during pregnancy to day 6.5 postconception, a significant increase in liver and spleen weights, and a significant increase in days to coitus--suggesting that a low level of toxicity was manifested. However, there were no adverse effects on reproductive performance with respect to conception and early embryonic development.

Developmental toxicity of cysteamine in the rat: effects on embryo-fetal development.

The reproductive and developmental safety of cysteamine has become an important issue to children with cystinosis because renal transplants and treatment with cysteamine reduce the complications associated with cystinosis and increase the lifespan of the affected children. In addition, there is the potential to decrease the severity or the incidence of renal Fanconi syndrome with administration of cysteamine to pregnant women carrying fetuses with cystinosis, and to ease significantly the burden of this disease throughout their lives. If cysteamine increases significantly the risk of fetal death, growth retardation or birth defects at doses used to treat women with cystinosis, treatment of the affected female should cease during pregnancy and would not be considered for fetal treatment. The goal of this study was to assess the developmental safety of exposure in utero to cysteamine in the rat. Pregnant rats were given cysteamine (as phosphocysteamine) from day 6.5 through day 18.5 postconception and fetuses were assessed for survival, growth, and structural abnormalities on day 20.5. Cysteamine was administered orally in doses of 0, 37.5, 75, 100, or 150 mg/kg/day. Cysteamine produced dose-dependent developmental toxicity with an apparent no adverse effect observed level of 75 mg/kg/day. Specific malformations were associated with this effect (cleft palate, kyphosis), as well as intrauterine growth retardation and fetal death at 100-150 mg/kg/day, without signs of maternal toxicity. Investigations continue into the mechanism for the developmental toxicity of cysteamine.

DSM-IV intermittent explosive disorder: a report of 27 cases.

BACKGROUND: The authors' objective was to provide data regarding the demographic, phenomenological, course of illness, associated psychiatric and medical comorbidity, family history, and psychiatric treatment response characteristics of rigorously diagnosed subjects who met DSM-IV criteria for intermittent explosive disorder. METHOD: Twenty-seven subjects meeting DSM-IV criteria for a current or past history of intermittent explosive disorder were given structured diagnostic interviews. The subjects' medical histories, family histories of psychiatric disorders, and responses to psychiatric treatments were also assessed. RESULTS: Most subjects described their intermittent explosive disorder symptoms as very distressing and/or highly problematic. All 27 subjects described aggressive impulses prior to their aggressive acts. Of 24 subjects who were systematically queried, 21 (88%) experienced tension with the impulses; 18 (75%), relief with the aggressive acts; and 11 (48%), pleasure with the acts. Most subjects stated that their aggressive impulses and acts were also associated with affective symptoms, particularly changes in mood and energy level. Twenty-five (93%) subjects had lifetime DSM-IV diagnoses of mood disorders; 13 (48%), substance use disorders; 13 (48%), anxiety disorders; 6 (22%), eating disorders; and 12 (44%), an impulse-control disorder other than intermittent explosive disorder. Subjects also displayed high rates of comorbid migraine headaches. First-degree relatives displayed high rates of mood, substance use, and impulse-control disorders. Twelve (60%) of 20 subjects receiving monotherapy with an antidepressant or a mood stabilizer reported moderate or marked reduction of their aggressive impulses and/or episodes. CONCLUSION: Intermittent explosive disorder appears to be a bona fide impulse-control disorder that may be related to mood disorder and may represent another form of affective spectrum disorder.

Uptake and processing of 59Fe-labelled and 125I-labelled rat transferrin by early organogenesis rat conceptuses in vitro.

The delivery of iron to the early organogenesis rat embryo has been studied, using 59Fe- and 125I-labelled rat transferrin. Rat conceptuses at 9.5 days postconception were cultured for 27 or 51 h in whole rat serum. Rat transferrin labelled with 59Fe was added for the final 0.1, 0.5, 6, 24 or 48 h of culture. Radioactivity accumulated progressively in both the embryo and the visceral yolk sac. Similar results were obtained when unconjugated 59Fe3+ was added to the rat serum used as culture medium. Both acid-soluble and acid-insoluble 59Fe were substantially present in the embryo and yolk sac after all exposure periods. When conceptuses were cultured in the presence of 125I-labelled rat transferrin, acid-soluble radioactivity was progressively released into the culture medium, but accumulation into the embryo and visceral yolk sac was slight and did not change with duration of exposure to the labelled protein. Similar findings were obtained using 125I-labelled bovine serum albumin. In these experiments, there was a close correspondence between the amount of iron accumulated by the embryo and visceral yolk sac in the final 24 h of a 51-h culture and the amount of transferrin converted into acid-soluble products in the same period. Visceral yolk sacs from 17.5-day pregnant rats were explanted and cultured in the presence of 59Fe-labelled rat transferrin, 125I-labelled rat transferrin or 125I-labelled bovine serum albumin, for periods up to 3 h. Again uptake of 59Fe increased with time of incubation, and the 125I-labelled proteins were digested to acid-soluble products which were released into the culture medium. The results indicate that transferrin delivers iron for incorporation into both the embryo and the visceral yolk sac, and are consistent with a mechanism involving receptor-mediated endocytosis of iron-laden transferrin by the cells of the visceral yolk sac. The transferrin itself appears to be quantitatively degraded, following delivery of iron to the yolk sac cells, a result that differs from findings in other cell types, in which the protein is not degraded but returns to the plasma membrane to participate in further cycles of iron acquisition and delivery.

Mechanisms of amino acid supply to the rat conceptus in normal and abnormal development.

Using in vitro and in vivo models, we have shown that protein is an important source of amino acids from 8.5 to 17.5 d postconception, a significant portion of postimplantation development in the rat. This satisfactorily explains why inhibition of protein pinocytosis and digestion by the visceral yolk sac may lead to congenital malformations, growth retardation, and intrauterine death during organogenesis and growth retardation and death during fetal stages. In humans, this process may be just as important and possibly just as vulnerable to inhibition, but other placental or paraplacental tissues may be involved in addition to the secondary yolk sac.

Investigations into mechanisms of amino acid supply to the rat embryo using whole-embryo culture.

The technique pioneered by D.A.T. New for the in vitro culture of early post-implantation rat embryos has been used to study nutritional mechanisms during early organogenesis. The results indicate that the principal route for amino acid supply to the 8.5- to 11.5-day embryo involves the endocytosis of proteins into cells of the visceral yolk sac endoderm, their digestion in lysosomes, and transmission of the amino acids to the growing embryo. Free amino acids constitute a comparatively unimportant source. Inhibition of either endocytosis or intralysosomal proteolysis diminishes amino acid supply to the embryo, and this can result in embryonic death or maldevelopment during organogenesis.

Leucine sources for the rat fetus.

Fetal and maternal plasma were assayed for the concentration of free leucine, acid-insoluble radioactivity and acid-soluble radioactivity at intervals after an intravenous bolus injection of [3H]leucine into anaesthetized pregnant rats at 17.5 days post-conception. The concentrations of total free leucine and of free [3H]leucine in maternal and fetal plasma were effectively unchanged from 5 to 180 min post-injection. Plasma free leucine concentrations in the fetus were five times those in the mother. The concentration of free [3H]leucine in fetal plasma was similar to that in maternal plasma. Thus the specific radioactivity of free leucine in fetal plasma is substantially lower than that in maternal plasma, indicating that a significant portion of the free leucine in plasma of the 17.5-day rat fetus comes from a source other than the free leucine in the maternal plasma. The data are consistent with a major contribution of amino acids coming from the degradation of extraembryonic protein in the yolk sac. Other possible sources of unlabelled leucine are discussed.

Leucine sources for 10.5-day rat conceptus in vivo.

Protein has been shown to be the principal source of leucine for the Day 8.5 to 10.5 rat conceptus in culture. It could be argued that this finding applies only after adaptation to culture conditions and does not apply in vivo. This possibility was investigated using an isotope-dilution technique after i.v. injection of [3H]leucine into Day 10.5 pregnant rats. Specific radioactivity of free leucine in the conceptus was 8 to 10% of that in maternal plasma. Slow exchange of leucine with the maternal circulation and fetal tissue protein turnover were judged to be inadequate as explanations for the observed isotope dilution. Taken together, our results and those from in vitro studies are consistent with a major contribution of leucine coming from the degradation of protein in vivo, probably involving the visceral yolk sac. Our results suggest that mechanisms of amino acid supply to the conceptus identified using whole-embryo culture mirror those in vivo.

Sources of amino acids for protein synthesis during early organogenesis in the rat. 3. Methionine incorporation.

Rat conceptuses at 9.5 days post-conception were cultured for 27 h in whole rat serum. [3H]Methionine, or rat serum proteins containing [3H]methionine, was introduced at 24 or 6 h before the termination of the culture. The total clearance of radioactivity into the embryo and the visceral yolk sac from the two sources was measured; also the extent to which the accumulated radioactivity was acid-insoluble. Similar experiments, but using [3H]leucine, were performed for comparison. The results indicate that free amino acid and protein can both serve as sources of amino acids for incorporation into the embryo and yolk sac, and it is estimated that in vivo over 95 per cent of the methionine (and the leucine) incorporated into these tissues arises from protein captured and digested by the yolk sac. Almost all the leucine accumulated into the conceptus is present as protein, but a larger fraction of the methionine accumulated is found in acid-soluble form. When the amino acids were delivered in the form of plasma proteins, the incorporation of methionine was two to three times more efficient than that of leucine, an observation most readily explained by leucine being provided in excess of requirements. In the light of reports that an adequate concentration of free methionine is important for the normal development of rat embryos in vitro, it is concluded that, although most of the amino acid required by the embryo is supplied as protein, the small fraction supplied as free amino acid may be critical for methionine but probably not for leucine.

Sources of amino acids for protein synthesis during early organogenesis in the rat. 4. Mechanisms before envelopment of the embryo by the yolk sac.

It was previously shown that uptake and digestion of protein by the visceral yolk sac supplies almost all of the amino acid needed by the 9.5-11.5-day rat conceptus cultured in vitro. Our aim was to test the hypothesis that protein uptake and digestion may not be as important as an amino acid source in the 8.5-9.5-day period, a stage of development before the yolk sac placenta envelops the embryo and before the vitelline circulation is established. Eight and a half-day rat conceptuses were cultured in serum supplemented with trace amounts of free [3H]leucine, [3H]leucine-containing serum proteins, free [3H]methionine or [3H]methionine-containing serum proteins. The incorporation of radiolabelled amino acid into acid-soluble and acid-insoluble fractions of the conceptus was determined. Leucine from either source was incorporated principally into proteins of the conceptus, but a greater proportion of the methionine incorporated was found in the low molecular weight fraction. It is estimated that 88 per cent of the leucine and 96 per cent of the methionine used by the conceptus was derived from protein in the culture serum; free amino acid comprised a minor supply source. We conclude that, despite the different anatomic relationships, the majority of amino acid incorporated into newly synthesized proteins of the conceptus very early in organogenesis is supplied by the digestion of protein in extraembryonic tissue, most likely the visceral yolk sac.

Is there a no-effect dose for corticosteroid-induced cleft palate? The contribution of endogenous corticosterone to the incidence of cleft palate in mice.

Teratology and genetic counselors are frequently asked whether very low exposures of drugs and chemicals can cause a child's congenital malformations. One critical factor on which the counseling is based is the dose. Because teratogenic effects follow a toxicologic dose-response curve with a no-effect dose, frequently counselors can refute a causal relationship because the dose was far below the no-observable-effect dose. Recently, some investigators have suggested that some teratogens which are present in physiologic levels such as cortisone, glucose, insulin, or sex steroids may contribute to the background incidence of congenital malformations and, therefore, there is no safe dose. Using corticosteroid-induced cleft palate in mice as the model, we conducted experiments to test this hypothesis. Adrenalectomy of A/J or CD-1 dams resulted in a reduction of endogenous corticosterone, but did not reduce the spontaneous incidence of cleft palate in the offspring. In A/J mice, the incidence of isolated cleft palate increased with adrenalectomy indicating that the spontaneous incidence of this defect is not due to endogenous corticosterone. Adrenalectomy did not affect the susceptibility of CD-1 mice to cortisone induced cleft palate demonstrating that endogenous corticosterone did not contribute significantly to the incidence of cleft palate induced by the exogenous corticosteroid. Finally, results in CD-1 mice clearly indicate that cortisone, like other teratogens, has a no-effect level for teratogenesis. These studies support the concept of a threshold in the dose-response relationship for corticosteroid-induced cleft palate in mice.

Pharmacokinetics of teratogenic antibodies administered to rats.

The serum levels of total and visceral yolk sac (VYS)-reactive sheep IgG have been determined following injection of a teratogenic sheep anti-VYS antiserum. After intravenous injection, levels of VYS-reactive IgG fell rapidly, with 75% of the amount in the injection removed in the first 5 min, and 90% by 60 min. By contrast, 90% or more of the total sheep IgG was still present at these times. A similar difference in clearance was seen after intraperitoneal injection, although the serum levels also reflected the presence of a pool of antiserum in the peritoneum and the simultaneous influx and efflux of IgG into and from the circulation. The clearance pattern was similar in pregnant and nonpregnant rats; it is concluded that antibodies against VYS-specific antigens comprise a very small fraction of VYS-reactive antibodies in the antiserum. ELISA and Western blot analysis indicated extensive cross-specificity with antigens present in rat tissues other than the VYS. Similar teratogenic effects were observed after intravenous or intraperitoneal injection of the antiserum at 8.5 days of gestation; we conclude that the proximal effect likely begins within the period immediately after the injection. The results are also considered within the context of published reports that the VYS shows structural and functional damage for several days after intraperitoneal administration of antiserum at 8.5 days.

The contribution of environmental teratogens to embryonic and fetal loss.

Environmental causes of human malformations account for approximately 10% of malformations, and less than 1% of all human malformations are related to prescription drug exposure, chemicals, or radiation. Malformations caused by drugs and other therapeutic agents are important, however, because these exposures are preventable. As we better understand the mechanisms of teratogenesis from all etiologies, we may learn how best to predict and test for teratogenicity and apply this knowledge to the prevention of human birth defects. Spontaneous abortions account for a much higher percentage of reproductive failure than do congenital malformations, but a very high proportion of aborted fetuses are malformed or defective. Therefore, the consequences of abortion are much different than the consequences of having a survival malformed infant, from both the medical and psychologic perspectives. We are less certain about the contribution of environmental, physical, and chemical agents to the incidence of spontaneous abortion. Although we realize that spontaneous abortions are a significant medical and emotional burden to a family, a surviving malformed infant can be greater burden to that family. We do not have an accurate picture of the contribution of environmental agents to the incidence of spontaneous abortion. At the present time, it would appear that only a very small proportion of abortions can be attributed to exposure to environmental toxicants during pregnancy (Tables 1 and 2). Conversely, it is the scientific and medical community's responsibility to prevent the introduction and use of agents that cause unwanted embryonic and fetal loss. Because the teratogenic and abortigenic effects of environmental agents differ, one cannot conclude that an agent is an abortifacient because it is teratogenic, nor can one conclude that an agent is not an abortifacient because it is not teratogenic (Table 2).

Effects of dose and dose protraction on embryotoxicity of 14.1 MeV neutron irradiation in rats.

The embryotoxic effects of neutron radiation on rodent embryos are documented, but there is disagreement about the dose-response relationship and the impact of protracting the dose. Pregnant rats were exposed to total absorbed doses of 0.15 to 1.50 Gy 14.1 MeV neutrons on day 9.5 after conception, coincident with the most sensitive stage of embryonic development for the induction of major congenital malformations. In general terms, the incidence of embryotoxic effects increased with increasing total absorbed dose. However, the dose-response relationship differed depending on the parameter of embryotoxicity chosen, namely, intrauterine death, malformations or very low body weight. In a second study, embryos were exposed to a single embryotoxic absorbed dose (0.75 Gy) administered at a range of dose rates, from 0.10 to 0.50 Gy/h. The results offer no evidence that protraction of this selected dose significantly increased or decreased the incidence or pattern of embryotoxicity of the neutron exposure used in this study. The results do not support the hypothesis of a linear dose-response relationship for the effects of prenatal neutron irradiation that contribute to embryotoxicity for total absorbed doses of 0.15 to 1.50 Gy.