Curcumin ameliorates streptozotocin-induced liver damage through modulation of endoplasmic reticulum stress-mediated apoptosis in diabetic rats.

We investigated the effect of curcumin on liver injury in diabetic rats induced by streptozotocin (STZ) through modulation of endoplasmic reticulum stress (ERS) and unfolded protein response (UPR). Experimental diabetes was induced by a single intraperitoneal injection of STZ (55 mg/kg), and curcumin was given at 100 mg/kg by gavage for 56 days. We observed that curcumin improved the morphological and histopathological changes, significantly decreased hepatic ERS marker protein: glucose-regulated protein 78, and improved liver function in diabetic rats. Moreover, treatment with curcumin markedly decreased the sub-arm of the UPR signaling protein such as phospho-double-stranded RNA-dependent protein kinase-like ER kinase, CCAAT/enhancer-binding protein homologous protein, tumor necrosis factor receptor-associated factor 2, and inositol-requiring enzyme1α; and inhibited tumor necrosis factor α, interleukin 1ß, phospho-p38 mitogen-activated protein kinase, and apoptosis signal-regulating kinase 1 in liver tissues of diabetic rats. Apoptotic and anti-apoptotic signaling proteins, such as cleaved caspase-3 and B-cell lymphoma 2, were significantly increased and decreased, respectively in diabetic rats; curcumin treatment prevented all of these alterations. In summary, our results indicate that curcumin has the potential to protect the diabetic liver by modulating hepatic ERS-mediated apoptosis, and provides a novel therapeutic strategy for the diabetic liver damage.

Zonisamide up-regulated the mRNAs encoding astrocytic anti-oxidative and neurotrophic factors.

Zonisamide has been proven as an effective drug for the recovery of degenerating dopaminergic neurons in the animal models of Parkinson's disease. However, several lines of evidence have questioned the neuroprotective capacity of zonisamide in animal models of Parkinson's disease. Although it suppresses dopaminergic neurodegeneration in animal models, the cellular and molecular mechanisms underlying the effectiveness of zonisamide are not fully understood. The current study demonstrates the effects of zonisamide on astrocyte cultures and two 6-hydroxydopamine-induced models of Parkinson's disease. Using primary astrocyte cultures, we showed that zonisamide up-regulated the expression of mRNA encoding mesencephalic astrocyte-derived neurotrophic factor, vascular endothelial growth factor, proliferating cell nuclear antigen, metallothionein-2, copper/zinc superoxide dismutase, and manganese superoxide dismutase. Similar responses to zonisamide were found in substantia nigra where the rats were pre-treated with 6-hydroxydopamine. Notably, pharmacological inhibition of 6-hydroxydopamine-induced toxicity by zonisamide pre-treatment was also confirmed using rat mesencephalic organotypic slice cultures of substantia nigra. In addition to this, zonisamide post-treatment also attenuated the nigral tyrosine hydroxylase-positive neuronal loss induced by 6-hydroxydopamine. Taken together, these studies demonstrate that zonisamide protected dopamine neurons in two Parkinson's disease models through a novel mechanism, namely increasing the expression of some important astrocyte-mediated neurotrophic and anti-oxidative factors.

Promoter CpG methylation in cancer cells contributes to the regulation of MUC4.

Mucin 4 (MUC4) is a high molecular weight transmembrane mucin that is overexpressed in many carcinomas and is a risk factor associated with a poor prognosis. In this study, we show that the DNA methylation pattern is intimately correlated with MUC4 expression in breast, lung, pancreas and colon cancer cell lines. We mapped the DNA methylation status of 94 CpG sites from -3622 to +29 using MassARRAY analysis that utilises base-specific cleavage of nucleic acids. MUC4-negative cancer cell lines and those with low MUC4 expression (eg, A427) were highly methylated near the transcriptional start site, whereas MUC4-positive cell lines (eg, NCI-H292) had low methylation levels. Moreover, 5-aza-2'-deoxycytidine and trichostatin A treatment of MUC4-negative cells or those with low MUC4 expression caused elevation of MUC4 mRNA. Our results suggest that DNA methylation in the 5' flanking region play an important role in MUC4 gene expression in carcinomas of various organs. An understanding of epigenetic changes in MUC4 may contribute to the diagnosis of carcinogenic risk and prediction of outcome in patients with cancer.

Plasma amantadine concentrations in patients with Parkinson's disease.

We determined plasma amantadine concentrations in patients with Parkinson's disease (PD) in daily clinical practice and investigated the relationship between plasma concentration and adverse reactions to clarify the safe therapeutic range. Seventy-eight consecutive PD patients on stable amantadine treatment were recruited. Plasma concentration of amantadine was measured 3h after the administration of morning amantadine dose. Serum creatinine was measured to estimate renal function. The mean daily dose of amantadine was 135.1+/-62.3mg/day, and the mean plasma amantadine concentration was 812.5+/-839.5 ng/ml (range, 91-4400 ng/ml). Plasma amantadine concentration increased according to increasing renal dysfunction. Three patients exhibited adverse reactions, such as myoclonus, hallucinations, and delirium, and all of them showed plasma amantadine concentration >3000 ng/ml. None of the three cases had previously shown such side effects. PD patients who have not developed any psychiatric symptoms as adverse reactions to the treatment may develop myoclonus, hallucination, or delirium when the plasma concentration of amantadine exceeds 3000 ng/ml. It is therefore recommended to use amantadine at the plasma concentration of less than 3000 ng/ml in the treatment of Parkinson's disease, especially in elderly patients.

Delayed L-DOPA-induced hyperalgesia.

Previously we reported on L-DOPA's antinociceptive effect on substance P-induced nociceptive behaviors in mice [Shimizu T, Iwata S, Morioka H, Masuyama T, Fukuda T, Nomoto M. Antinociceptive mechanism of L-DOPA. Pain 2004;110;246-9.]. Since significant hyperalgesia was noted following antinociception, our study was designed to investigate the mechanism of this hyperalgesia. Nociceptive behaviors were enhanced 2 h after L-DOPA administration. L-DOPA induced hyperalgesia occurred after conversion to dopamine because co-administration of benserazide, a DOPA decarboxylase inhibitor, completely abolished the L-DOPA-induced hyperalgesia. The D2 receptor agonist, quinpirole, depressed these behaviors entirely, while the D1 antagonist, SCH23390, inhibited the enhancement of these behaviors by L-DOPA. The D2 receptor antagonist, sulpiride, which induced hyperalgesia of the substance P-induced behaviors in naive mice, did not have any effects on L-DOPA-induced hyperalgesia. Spinal cord dopamine content increased rapidly after L-DOPA administration, exhibiting levels 100 times greater than baseline, and then returned to control after 1 h. These results suggested that the dopaminergic inhibitory system for pain sensation was temporarily impaired by excess amounts of exogenous dopamine that were derived from L-DOPA and both D1 and D2 receptors were involved in L-DOPA-induced hyperalgesia.

Expression of mucins (MUC1, MUC2, MUC5AC and MUC6) in mucinous carcinoma of the breast: comparison with invasive ductal carcinoma.

AIMS: Mucinous carcinoma of the breast usually shows less frequent lymph node metastasis and more favourable outcome compared with invasive ductal carcinoma. The aim of this study is to compare the expression profiles of several mucins in mucinous carcinomas and invasive ductal carcinomas to gain insight into the relationship between the less aggressive biological nature of mucinous carcinoma and the role of mucins. METHODS AND RESULTS: We examined the expression profiles of MUC1 (membrane-bound mucin) of different glycoforms (from non-glycosylated form to fully glycosylated form), MUC2 (intestinal type secretory mucin), MUC5AC (gastric surface type secretory mucin) and MUC6 (gastric pyloric gland type secretory mucin) in 17 mucinous carcinomas and 46 invasive ductal carcinomas using immunohistochemistry. Various glycoforms of MUC1 were expressed frequently in both mucinous carcinomas (65-100%) and invasive ductal carcinomas (92-100%), although non-glycosylated MUC1 (MUC1/CORE) and fully glycosylated MUC1 (MUC1/HMFG-1) showed significantly lower expression rates in mucinous carcinomas compared with those in invasive ductal carcinomas. The expression rates of MUC2 (94%) and MUC6 (71%) in mucinous carcinomas were significantly higher than those of MUC2 (15%) and MUC6 (15%) in invasive ductal carcinomas. There was no significant difference in the expression rate of MUC5AC in mucinous carcinomas (12%) and that in invasive ductal carcinomas (4%). CONCLUSIONS: The expression rate of MUC1/CORE and MUC1/HMFG-1, which is related to poor prognosis in the gastric and colorectal cancers, is low in mucinous carcinomas. The high expression rate of gel-forming secretory mucins (MUC2 and MUC6) in mucinous carcinoma suggests that high production of these types of mucins may act as a barrier to cancerous extension resulting in their less aggressive biological behaviour.

Circulating immunoglobulin-bound transforming growth factor beta at a late tumour-bearing stage impairs antigen-specific responses of CD4+ T cells.

In order to elucidate the mechanisms by which tumour-specific CD4+ T-cell responses are impaired during tumour development, an attempt was made to identify factors which impair CD4+ T-cell responses at a late tumour-bearing stage. Plasma from mice bearing B16 melanoma for 30 days (plasma d30) showed a more profound immunosuppressive effect on the in vitro proliferation of unrelated antigen-specific CD4+ T cells in the presence of both antigen and antigen-presenting cells (APC) than plasma from naïve mice. The level of plasma transforming growth factor (TGF)- was elevated in mice bearing B16 melanoma for 30 days compared with naïve mice, and the suppressive effect of plasma d30 was partially diminished by the neutralization of TGF-. Interestingly, immunoglobulin (IgG)-bound TGF-, but not IgG-unbound TGF-, in plasma d30 was suggested to be responsible for the immunosuppressive activity. In addition, no suppressive effect of plasma d30 was observed when antigen was added as a class II peptide, thus suggesting that the impaired proliferation of CD4+ T cells in the presence of plasma d30 was due to a dysfunction of antigen uptake/processing by APC. Furthermore, dissociation between IgG and TGF- resulted in a loss of the suppressive activity of plasma d30. Taken together, these results suggest that circulating IgG-bound TGF- is, at least in part, responsible for the impaired responses of CD4+ T cells at the late tumour-bearing stage by suppressing antigen uptake/ processing by APC.

Chronic estrogen treatment replaces striatal dopaminergic function in ovariectomized rats.

Eight-week-old female Sprague-Dawley rats were divided into three groups: ovariectomized rats (OVX); ovariectomized rats treated with estradiol valerate (E2), 20 microg subcutaneously (s.c.) twice weekly for 12 weeks (OVX+E2 group); and sham-operated control rats treated with vehicle alone (controls). Spontaneous locomotor activity was measured for 24 h, and then again after the administration of methamphetamine (1 mg/kg, i.p.). In addition, striatal contents of dopamine (DA) and its metabolites were measured. Using an in vivo microdialysis technique, changes in extracellular striatal dopamine concentration were studied in a separate set of similarly treated rats after the administration of methamphetamine (0.2 mg/kg, i.p.). Spontaneous locomotor activity decreased in the OVX group, and estradiol replacement reversed this decreased activity. No significant differences were observed in the contents of DA and its metabolites at the striatum among the three groups. The basal output of DA at the striatum was lower in the OVX group than in those of the other two groups. Extracellular DA concentration following methamphetamine administration was also lower in the rats of OVX group. These results indicate that ovariectomy decreases spontaneous locomotor activity, response to methamphetamine, and striatal DA release in the female rats. Chronic replacement of estrogen reversed spontaneous locomotor activity and DA release by the striatum. These results suggest that chronic administration of estrogen may be beneficial in the treatment of female menopausal patients with Parkinson's disease.

[Pharmacological treatments of Parkinson's disease].

Antiparkinsonian agents applied or under the investigation for the treatment of patients with Parkinson's disease were reviewed. Tremor, akinesia, rigidity and postual instability are key signs of Parkinson's disease. The most important one is akinesia, which includes decreased spontaneous locomotor activity, slowness of movement, awkwardness and freezing. The main pathophysiology of Parkinson's disease is neurodegeneration of nigrostriatal dopaminergic neurons. Neurotoxins or oxidative stress to the dopaminergic neurons have been discussed as one of the etiologies of degeneration. Antioxidant or neuroprotective agents will be the future drugs for Parkinson's disease. At present, supplement of dopamine by levodopa administration, retarding the metabolism of levodopa or dopamine by a dopa decarboxylase inhibitor (DCI), MAO-B (monoamine oxidase inhibitor type B) inhibitor or catechol-O-methyltransferase (COMT) inhibitor, dopamine receptor agonists, anticholinergic agents, dopamine release enhancer/uptake inhibitor, N-methyl-D-aspartate (NMDA) receptor antagonists are applied for the treatment of Parkinson's disease. New agents such as adenosine receptor antagonists, serotonergic agents and nicotinic receptor agonists are under investigation. Agents to facilitate the growth of nerves or to inhibit degeneration of nerves are also studied and will be developed for the treatment of Parkinson's disease in the future. In the case of familial Parkinson's disease, abnormal genes were identified. Gene therapy might be another future treatment for these cases.

Transcriptional activation of the human HMG1 gene in cisplatin-resistant human cancer cells.

The nonhistone chromosomal protein, high mobility group 1 (HMG1), which is ubiquitously expressed in higher eukaryotic cells, preferentially binds to cisplatin-modified DNA. The observation that HMG1 is overexpressed in cisplatin-resistant human cancer cells suggests that cisplatin resistance may be closely associated with HMG1. To decipher the mechanism of HMG1 overexpression in cisplatin-resistant cells, we isolated two overlapping genomic DNA clones containing the entire human HMG1 gene. These clones, which span approximately 15 kb of contiguous DNA, include 5 kb of the 5' flanking region as well as the entire coding sequence. We sequenced 1500 bp upstream of the first exon. The segment proximal to the transcription initiation site did not contain a TATA box but did possess an activating transcription factor site, an activator protein-2 site, one CCAAT box, and two CCAAT-binding transcription factor/nuclear factor-1 (CTF/NF-1) sites. HMG1 promoter activity was 3-10-fold higher in cisplatin-resistant KB-CP20 cells than in parental KB cells. An in vivo footprint experiment showed several differences of dimethyl sulfate modifications between KB and KB-CP20 cells in the area around the CTF/NF-1 sites. In addition, electrophoretic gel mobility shift assays showed that binding of a nuclear factor from cisplatin-resistant cells to the CTF/NF-1 site was significantly higher than the binding of the same factor from parental cells. Semiquantitative reverse transcription-PCR and Western blot analysis also showed that expression of CTF/NF-1 was 3-20-fold higher in the resistant cell line than in its parental counterpart. These findings suggest that, in cisplatin-resistant cells, the expression of HMG1 gene product is enhanced at the transcriptional level and that this probably occurs through the enhanced expression of the CCAAT binding factor, CTF/NF-1.

Y box-binding protein-1 binds preferentially to single-stranded nucleic acids and exhibits 3'-->5' exonuclease activity.

We have previously shown that Y box-binding protein-1 (YB-1) binds preferentially to cisplatin-modified Y box sequences. Based on structural and biochemical data, we predicted that this protein binds single-stranded nucleic acids. In the present study we confirmed the prediction and also discovered some unexpected functional features of YB-1. We found that the cold shock domain of the protein is necessary but not sufficient for double-stranded DNA binding while the C-tail domain interacts with both single-stranded DNA and RNA independently of the cold shock domain. In an in vitro translation system the C-tail domain of the protein inhibited translation but the cold shock domain did not. Both in vitro pull-down and in vivo co-immunoprecipitation assays revealed that YB-1 can form a homodimer. Deletion analysis mapped the C-tail domain of the protein as the region of homodimerization. We also characterized an intrinsic 3'-->5' DNA exonuclease activity of the protein. The region between residues 51 and 205 of its 324-amino acid extent is required for full exonuclease activity. Our findings suggest that YB-1 functions in regulating DNA/RNA transactions and that these actions involve different domains.

Mucosal damage and recovery of the intestine after prolonged preservation and transplantation in dogs.

BACKGROUND: Although much is known about the mucosal damage that occurs after intestinal warm ischemia and reperfusion and its recovery, little is known about the effect of cold preservation and transplantation on the mucosa. We studied the electrophysiological, biochemical, and histological changes of the intestinal mucosa after preservation for 24 hr and subsequent transplantation. METHODS: The small intestines from adult mongrel dogs were harvested. The intestines were orthotopically autotransplanted immediately (control group) or after preservation for 24 hr (preservation group). Jejunal and ileal tissues were taken before harvesting, at the end of preservation, 1 hr after reperfusion, and on postoperative days 3, 7, 14, and 28. The Ussing chamber method was used to study the electrophysiologic changes. Tissue maltase, diamine oxidase, and ornithine decarboxylase were measured. A histological analysis was also performed. RESULTS: Control group grafts showed no evident deterioration in electrophysiology, biochemistry, or morphology. In contrast, preservation group grafts exhibited electrophysiological and biochemical degradation, complete denudation of the villi, and crypt injury (especially in the ileum) after reperfusion. Electrophysiologic function and the mucosa biochemical marker recovered within 3 days in the jejunum and within 7-14 days in the ileum; however, histological recovery of mucosal injury required 28 days in the jejunum and more than 28 days in the ileum. CONCLUSIONS: Our study showed that despite severe destruction of mucosal integrity by prolonged preservation and transplantation, the intestinal mucosa has an enormous regenerative capacity. Our study also showed that regeneration was more pronounced in the jejunum than in the ileum.

Persistent infection with Listeria monocytogenes in the kidney induces anti-inflammatory invariant fetal-type gammadelta T cells.

After intraperitoneal inoculation with Listeria monocytogenes, gammadelta T cells appear in the peritoneal cavity preceding the appearance of alphabeta T cells. Such gammadelta T cells predominantly express T-cell receptor (TCR)Vgamma1/Vdelta6, develop through an extrathymic pathway, and contribute to host defence against the bacteria. We have observed a gradual increase in gammadelta T cells in kidneys of mice after intrarenal inoculation with L. monocytogenes, which resulted in an unusually long-lasting local infection. In this study, we examined the characteristics and the roles of the gammadelta T cells induced in this model. It was found that these gammadelta T cells predominantly expressed TCRVgamma6/Vdelta1 with canonical junctional sequences identical to those expressed on fetal thymocytes. Although depletion of such gammadelta T cells in vivo did not affect the number of bacteria, it resulted in histologically exacerbated inflammation in the kidneys. These results indicate that a persistent infection with L. monocytogenes in kidneys induces a different kind of gammadelta T cell from that induced after intraperitoneal infection. The former expresses invariant fetal-type Vgamma6/Vdelta1+TCR and plays a regulatory role in resolution of inflammation.

Interaction with p53 enhances binding of cisplatin-modified DNA by high mobility group 1 protein.

A nonhistone chromosomal protein, high mobility group (HMG) 1, is ubiquitous in higher eukaryotic cells and binds preferentially to cisplatin-modified DNA. HMG1 also functions as a coactivator of p53, a tumor suppressor protein. We investigated physical interactions between HMG1 and p53 and the influence of p53 on the ability of HMG1 to recognize damaged DNA. Using immunochemical coprecipitation, we observed binding of HMG1 and p53. Interaction between HMG1 and p53 required the HMG A box of HMG1 and amino acids 363-376 of p53. Cisplatin-modified DNA binding by HMG1 was significantly enhanced by p53. An HMG1-specific antibody that recognized the A box of this protein also stimulated cisplatin-modified DNA binding. These data suggest that an interaction with either p53 or antibody may induce conformational change in the HMG1 A box that optimizes DNA binding by HMG1. Interaction of p53 with HMG1 after DNA damage may promote activation of specific HMG1 binding to damaged DNA in vivo and provide a molecular link between DNA damage and p53-mediated DNA repair.

Regulation of GAP-43 protein and mRNA in nigrostriatal dopaminergic neurons after the partial destruction of dopaminergic terminals with intrastriatal 6-hydroxydopamine.

Changes in the level of GAP-43 and its mRNA in nigrostriatal dopaminergic neurons in an animal model of the presymptomatic period of Parkinson's disease were measured to find the characteristic features of GAP-43 in nigrostriatal dopaminergic neurons. Since the dopaminergic neurons possess a relatively large amount of GAP-43 protein and mRNA, the dopaminergic neurons must be endowed with specific functions related to those of GAP-43. In this study, dopaminergic axon terminals were partially destroyed by intrastriatal 6-hydroxydopamine (6-OHDA). Rats were decapitated 3, 14, and 56 days following treatment. Levels of GAP-43 and tyrosine hydroxylase (TH) in the striatum were detected by immunoblotting and quantified. The number of GAP-43 mRNA-positive neurons and that of TH mRNA-positive neurons in the substantia nigra pars compacta (SNc) were detected by in situ hybridization using alkaline phosphatase (ALP)-labeled probes. Levels of GAP-43 in the striatum showed no significant alteration during the period of the experiment, although levels of TH were gradually restored. The number of GAP-43 mRNA-positive neurons as well as that of TH mRNA-positive neurons in the SNc decreased. These results suggests that dopaminergic neurons restore their axon terminals with little change in GAP-43, and that transcription and/or stability of GAP-43 mRNA in the dopaminergic neurons are susceptible to the toxin, although the dopaminergic neurons can maintain the translational product in the terminals. This feature may be related with a degeneration of dopaminergic neurons in Parkinson's disease.

[Functions and metabolic rates of transmitters in the basal ganglia].

The caudate nucleus and putamen showed different neuronal circuits with the cerebral cortex and different functions in motor control. The dopamine content was same in the caudate nucleus and putamen, however, the turnover ratio was higher in the putamen than in the caudate nucleus, when it was studied in the control or MPTP-treated marmosets. The dopaminergic neurons projecting to the putamen seemed to show higher activity than those projecting to the caudate nucleus. The caudate nucleus is long in the shape and the dopamine turnover ratio was higher in the former part than in the latter part in normal marmosets. Administration of fluvoxamine increased akinesia in MPTP-treated common marmosets, and systemic administration of 8-OHDPAT, a 5-HT1A receptor or autoreceptor agonist reversed parkinsonism in MPTP-treated common marmosets. These results indicated that serotonergic neurons potentiated hypoactivity in parkinsonism and suppression of serotonergic function may contribute to the remission of parkinsonism.

The metabolic rate and vulnerability of dopaminergic neurons, and adenosine dynamics in the cerebral cortex, nucleus accumbens, caudate nucleus, and putamen of the common marmoset.

The pathophysiology of the striatum and cerebral cortex were studied from the pharmacological aspect. Investigation of the dopamine content in the cerebral cortex revealed that the premotor and motor area showed the highest level (61+/-6.2 ng/g). Intravenous injection of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) at a dose of 10 mg/kg reduced the dopamine content in the caudate nucleus and putamen to 2-3% of the control level in common marmosets, while it fell to 60% in the nucleus accumbens. There was no alteration of the dopamine content in the cerebral cortex. Immunohistochemical staining for tyrosine hydroxylase in the midbrains of MPTP-treated marmosets showed almost complete disappearance of dopaminergic cells from the substantia nigra and good preservation of cells in the ventrotegmental area. Dopaminergic cells projecting to the caudate/putamen, nucleus accumbens, and cerebral cortex showed marked, moderate, and no vulnerability to MPTP, respectively. After systemic administration of MPTP, dopaminergic neurons projecting to the caudate nucleus and putamen were damaged equally. However, the compensatory increase of dopamine turnover was more prominent in the putamen than in the caudate nucleus. Thus, nigroputaminal dopaminergic neurons may have a higher level of activity than neurons in the caudate. The neural connections and functions of the caudate nucleus and putamen have already been differentiated anatomically or physiologically. This compensatory increase of the dopamine turnover rate is another aspect of functional differences between the caudate nucleus and putamen. Investigation of the dopamine content in the head, body, and tail of the caudate nucleus showed no differences in the concentration of dopamine. However, a study of the metabolic rate of dopamine using alpha-methyl-p-tyrosine, a tyrosine hydoxylase inhibitor, showed higher metabolism of dopamine in the head of the caudate nucleus in common marmosets. Thus, dopaminergic neurons projecting to the caudate nucleus may show topographical differences in their firing rates. A microdialysis study indicated an increase in the metabolism of adenosine in the striatum of MPTP-treated animals. Cholinergic neurons are interneurons and are one of the main sources of adenosine in the striatum. Dopaminergic input from the substantia nigra acting on cholinergic neurons was decreased following MPTP treatment. The increase of adenosine metabolism suggested that cholinergic neurons in the striatum receive inhibitory inputs from nigrostriatal dopaminergic neurons.

Expression of UDP-GalNAc: polypeptide N-acetylgalactosaminyltransferase isozymes T1 and T2 in human colorectal cancer.

Uridine diphosphate (UDP)-GalNAc: polypeptide N-acetylgalactosaminyltransferase (GalNAc transferase) catalyzes the initial step in mucin type O-glycosylation, and its expression has been assumed to be altered between normal epithelial cells and cancer cells. We studied the alteration of GalNAc transferase expression during the carcinogenesis of human colorectal epithelial cells. We produced polyclonal antibodies against synthetic polypeptides with specific sequence to two GalNAc transferase isozymes, T1 and T2. Surgically resected specimens from 50 patients with colorectal cancer were immunohistochemically stained, and the staining grade (percentage of positively stained cells) was compared between cancer and its normal counterpart in the same specimen. Significant signals for both T1 and T2 expression were seen in the supranuclear region of normal and cancer cells, indicating the subcellular localization of the enzymes in the Golgi apparatus. The prevalence of positive staining for T1 and T2 expression in colorectal cancer was significantly higher than that in normal epithelium (P < 0.05). However, the difference in staining grades between cancer and normal tissues varied in each patient. These results indicate that there is variability in the expression patterns of GalNAc transferase isozymes in normal and cancerous cells colorectal among individuals.

TH protein and mRNA in nigrostriatal dopaminergic neurons are down-regulated by continuous but not intermittent apomorphine.

Levels of tyrosine hydroxylase (TH) and TH mRNA were measured after administration of dopamine agonists for a long period of time to elucidate the long-term feedback inhibition of dopamine synthesis in nigrostriatal dopaminergic neurons. Continuous infusion, which desensitized presynaptic dopamine receptors, but not repeated administration, down-regulated TH and TH mRNA levels. This suggests levels of TH protein and mRNA are only feedback inhibited by the continuous stimulation of postsynaptic dopamine receptors.