Interspecies differences in the kinetic properties of deoxycytidine kinase elucidate the poor utility of a phase I pharmacologically directed dose-escalation concept for 2-chloro-2′-deoxyadenosine

2-Chloro-2-deoxyadenosine (CdA, Cladribine), is a purine antimetabolite currently under investigation in phase II clinical trials for the treatment of lymphoid malignancies. Significant differences in CdA toxicity between mice and humans were observed during phase I clinical evaluation. For the elucidation of interspecies differences in drug toxicity the pharmacokinetics of CdA after subcutaneous injection and the kinetic properties of the CdA-phosphorylating enzyme, deoxycytidine kinase (dCK), were compared in mice and humans. The ratio of the dose lethal to 10% of mice (LD₁₀) to the maximum tolerated dose (MTD) in humans was 50 and the ratio of the area under the curve obtained at approximately one-half the LD₁₀ (AUC_{approx. one-half the LD 10})/AUC_MTD was 49. A significant interspecies difference was observed in the kinetic properties of dCK, the main CdA-activating enzyme. With CdA as a substrate, the Michaelis constant (K _m) of dCK in crude extracts of mouse thymus was 10 times higher than that in human thymus. An approximately 9-fold interspecies difference in maximum velocity (V_max)/K _m indicated a higher efficiency of dCK for CdA in humans than in mice. The peak plasma concentration was 210 times higher and exceeded theK _m in mice. Initial and terminal half-lives were approximately 7 times shorter in mice and trough levels were similar in mice and humans. Thus, the differences in AUCs at equitoxic doses are largely explained by differences in the target enzyme properties and the pharmacokinetic pattern. The observed lower tolerance for CdA in humans as compared with mice confirms the view that antimetabolites may not be good candidates for pharmacokinetically guided dose-escalation schemes unless detailed information on interspecies variability in drug bioactivation is available.     

Developmental damage, increased lipid peroxidation, diminished cyclooxygenase-2 gene expression, and lowered prostaglandin E2 levels in rat embryos exposed to a diabetic environment

Previous experimental studies suggest that diabetic embryopathy is associated with an excess of radical oxygen species (ROS), as well as with a disturbance of prostaglandin (PG) metabolism. We aimed to investigate the relationship between these pathways and used hyperglycemia in vitro (embryo culture for 24-48 h) and maternal diabetes in vivo to affect embryonic development. Subsequently, we assessed lipid peroxidation and gene expression of cyclooxygenase (COX)-1 and -2 and measured the concentration of prostaglandin E2 (PGE2) in embryos and membranes. Both hyperglycemia in vitro and maternal diabetes in vivo caused embryonic dysmorphogenesis and increased embryonic levels of 8-epi-PGF2alpha, an indicator of lipid peroxidation. Addition of N-acetylcysteine (NAC) to the culture medium normalized the morphology and 8-epi-PGF2alpha concentration of the embryos exposed to high glucose. Neither hyperglycemia nor diabetes altered COX-1 expression, but embryonic COX-2 expression was diminished on gestational day 10. The PGE2 concentration of day 10 embryos and membranes was decreased after exposure to high glucose in vitro or diabetes in vivo. In vitro addition of NAC to high glucose cultures largely rectified morphology and restored PGE2 concentration, but without normalizing the COX-2 expression in embryos and membranes. Hyperglycemia/diabetes-induced downregulation of embryonic COX-2 gene expression may be a primary event in diabetic embryopathy, leading to lowered PGE2 levels and dysmorphogenesis. Antioxidant treatment does not prevent the decrease in COX-2 mRNA levels but restores PGE2 concentrations, suggesting that diabetes-induced oxidative stress aggravates the loss of COX-2 activity. This may explain in part the antiteratogenic effect of antioxidant treatment.     

Somatostatin receptor scintigraphy during treatment with lanreotide in patients with neuroendocrine tumors

To investigate possible changes in somatostatin receptor expression during treatment with high dose lanreotide, eight patients with neuroendocrine tumors were investigated by [¹¹¹In-DTPA-D-Phe¹]-octreotide scintigraphy before and during treatment. The spleen-to-background ratio decreased in all patients, whereas tumor-to-background ratio revealed a heterogeneous pattern with an average increase of 50% (−79% to +1,087%). This finding indicates that lanreotide treatment may influence the binding of radioactively labeled somatostatin to the spleen, while changes in the binding to functioning somatostatin receptors in tumor cells are more complex and not clearly related to treatment.     

Male Reduction Mammaplasty After Vertical Banded Gastroplasty

Background: Breast reduction surgery is common in females; however, in males it is mainly due to gynecomastia. After weight reduction following obesity surgery, it is a problem in women, but also in some men. Method: One patient is described in whom the weight reduction declined from BMI 52 to BMI 36 after vertical banded gastroplasty, giving the patient ptotic breasts. Results: The patient underwent reduction mammaplasty with lateral single-based cutaneous flaps, and a total of 1,000 g was removed. Conclusion: Reduction mammoplasty can be performed in males with the methods used today, after successful weight loss following obesity surgery.     

Diverse changes in fibre type composition of the human lateral pterygoid and digastric muscles during aging

The fibre type composition of the superior and inferior portions of the human lateral pterygoid and the anterior and posterior bellies of the digastric muscles of five elderly subjects (mean age 73 years) was studied by morphological and enzyme-histochemical methods. Both muscles showed significant age-related changes in fibre type composition as compared with previous data for young adults. In the lateral pterygoid we observed a large proportion of type IIA fibres, which are rare or absent in young adults, and muscle fibre atrophy and an increased variability in fibre diameter. The digastric muscle of elderly showed a decrease in the proportion of type IIB fibres. The only difference in age-related changes between muscle portions was found in the lateral pterygoid with fibre atrophy in its inferior portion. Both the lateral pterygoid and digastric muscles are known to be active in mandibular depression (jaw opening) and horizontal positioning of the mandible. The present results and previous data from young adults show that the lateral pterygoid and digastric muscles differ not only in fibre type composition, but also in muscular changes following aging. This suggests that, even if they are simultaneously active, they fulfill different, specific tasks in natural jaw function. The differences in age-related changes in fibre type composition between these two muscles indicate that mechanisms underlying their alterations during aging are muscle specific. The results indicate that, although nerve supply and developmental history are essential for fibre composition of skeletal muscles, functional tasks and demands are of major importance.     

Isolation of sex-specific cDNAs from fetal mouse brain using mRNA differential display and representational difference analysis

Comparing female and male brain structures reveals a variety of sex differences in many vertebrates. These differences are manifested throughout the brain, in regions such as the hypothalamus, the preoptic area and the amygdala. Some are thought to be induced during the fetal period by the effect of steroid hormones produced in the gonads. It is well-established that fetal androgens, probably through the conversion to estrogen by the enzyme aromatase, masculinize the nervous system and set adult mounting behavior in rodents. However, less is known about molecular mechanisms involved in gender-specific development of the brain. We have taken a broad approach to isolate sex-specific genes from fetal brain. mRNAs from 18.5 days post-coitum (dpc) female and male mouse brain were screened with the classical and the recently developed signal peptide differential display (SPDD) and with representational difference analysis of cDNA (cDNA-RDA). Two sex-specific cDNAs were isolated, F29 and M17, corresponding to the female-specific Xist gene and the male-specific Smcy gene, respectively.     

Delta-opioid receptor immunoreactivity on astrocytes is upregulated during mitosis

Endogenous opioid peptides and opioid receptors are expressed by brain cells early during normal development, and exogenous opiate exposure in this period is known to affect brain cell proliferation and maturation. Despite the abundant evidence that opioids affect brain development, little is known about the mechanisms involved. In this study cortical astrocytes in primary culture were examined immunohistochemically by using antibodies against the opioid receptors. The immunoreactivity for delta-opioid receptors was strongly upregulated during mitosis with an increase in immunostaining that started in early prophase and lasted through the M-phase to cytokinesis. Similar effects could not be observed when antibodies against the mu- or kappa-opioid receptor subtypes were used. Cultured neurons and microglia presented a strong and homogenous immunostaining for the delta-opioid receptor and no further upregulation of immunoreactivity could be detected in these cells. The presence of functional delta-opioid receptors on the mitotic astrocytes was verified by using microspectrofluorometry for detection of delta-opioid agonist induced changes in intracellular free calcium concentrations ([Ca2+]i). In these experiments fluo-3/AM incubated cells showed a rapidly induced delta-opioid agonist (DPDPE, 10(-6) M) evoked increase in [Ca2+]i. These results suggest an upregulation of the delta-opioid receptors that could represent a mechanism involved in the response to opioids in the developing brain.     

Analysis of CSF amino acids in young patients with generalised refractory epilepsy during an add-on study with lamotrigine

The effect of add-on administration of lamotrigine (1-12 mg/kg per day, 2-12 months) on the levels of neurotransmission related amino acids including gamma-aminobutyric acid (GABA), glutamate, aspartate, glycine and antiepileptic drugs (AEDs) in lumbar cerebrospinal fluid (CSF) was studied in 22 children and young adults with generalised therapy resistant epilepsy. Two lumbar punctures were performed, one prior to, and one following a mean of 5 months (2-12 months) of lamotrigine treatment. Lamotrigine decreased seizure incidence and severity in 12 of the 22 patients without influencing CSF GABA, glutamate, aspartate or glycine levels. Lamotrigine did not alter the concentrations of AEDs in CSF or plasma. However, CSF GABA levels were 86% higher in those patients also treated with gamma-vinyl-GABA (vigabatrin, GVG) compared with patients treated with other combinations and this was not altered by co-medication with lamotrigine. The proposed mechanism of action of lamotrigine, namely that it may inhibit glutamate release in the CNS, is not reflected by changes in CSF glutamate levels. The present findings indicate that CSF GABA, glutamate, aspartate and glycine levels may not be useful as in vivo neurochemical markers in young patients responding to the therapeutic dose of lamotrigine used in this study.     

Developmental neurotoxicity of four ortho-substituted polychlorinated biphenyls in the neonatal mouse

The objective of the present study was to investigate whether neonatal exposure to single PCB (polychlorinated biphenyl) congeners 2,4,4'-trichlorobiphenyl (IUPAC 28), 2,2',5,5'-tetrachlorobiphenyl (IUPAC 52), 2,3',4,4',5-pentachlorobiphenyl (IUPAC 118) and 2,3,3',4,4',5-hexachlorobiphenyl (IUPAC 156) when given as one single dose (0.7-14 μmol/kg body weight per os) to 10-day-old male NMRI mice could induce persistent neurotoxic effects in the adult animal. Furthermore, to ascertain whether behavioural aberrations, both in spontaneous behaviour and in learning and memory function, were followed by changes in the cholinergic and/or the dopaminergic system. It was found that neonatal exposure to lightly chlorinated ortho-substituted PCBs, 2,4,4'-tri- and 2,2',5,5'-tetrachlorobiphenyls, can induce persistent aberrations in spontaneous behaviour. Neonatal exposure to 2,2',5,5'-tetrachlorobiphenyl also affected learning and memory functions in the adult animal. In the animals showing a deficit in memory and learning function, the cholinergic nicotinic receptors in the cerebral cortex were affected. Exposure to 2,3',4,4',5-penta- and 2,3,3',4,4',5-hexachlorobiphenyl, mono-ortho congeners ('co-planar-like'), in the same dose range did not cause any significant change in the investigated behavioural variables, spontaneous and swim-maze behaviour.