Construction of novel immune-related signature for prediction of pathological complete response to neoadjuvant chemotherapy in human breast cancer.

BACKGROUND: The aim of this study was to construct a novel prediction model for the pathological complete response (pCR) to neoadjuvant chemotherapy (NAC) using immune-related gene expression data. PATIENTS AND METHODS: DNA microarray data were used to perform a gene expression analysis of tumor samples obtained before NAC from 117 primary breast cancer patients. The samples were randomly divided into the training (n = 58) and the internal validation (n = 59) sets that were used to construct the prediction model for pCR. The model was further validated using an external validation set consisting of 901 patients treated with NAC from six public datasets. RESULTS: The training set was used to construct an immune-related 23-gene signature for NAC (IRSN-23) that is capable of classifying the patients as either genomically predicted responders (Gp-R) or non-responders (Gp-NR). IRSN-23 was first validated using an internal validation set, and the results showed that the pCR rate for Gp-R was significantly higher than that obtained for Gp-NR (38 versus 0%, P = 1.04E-04). The model was then tested using an external validation set, and this analysis showed that the pCR rate for Gp-R was also significantly higher (40 versus 11%, P = 4.98E-23). IRSN-23 predicted pCR regardless of the intrinsic subtypes (PAM50) and chemotherapeutic regimens, and a multivariate analysis showed that IRSN-23 was the most important predictor of pCR (odds ratio = 4.6; 95% confidence interval = 2.7-7.7; P = 8.25E-09). CONCLUSION: The novel prediction model (IRSN-23) constructed with immune-related genes can predict pCR independently of the intrinsic subtypes and chemotherapeutic regimens.

Recurrence risk score based on the specific activity of CDK1 and CDK2 predicts response to neoadjuvant paclitaxel followed by 5-fluorouracil, epirubicin and cyclophosphamide in breast cancers.

BACKGROUND: We established the cell cycle profiling (C2P) assay for specific activity (SA; activity/expression) of cyclin-dependent kinases (CDKs). C2P risk score (C2P-RS) based on CDK1 and CDK2 SAs was significantly associated with relapse in breast cancer (BC). This study was conducted to investigate the predictive value of C2P-RS for neoadjuvant chemotherapy (NAC). PATIENTS AND METHODS: Among 124 eligible patients, 122 were treated with weekly paclitaxel followed by 5-fluorouracil, epirubicin and cyclophosphamide (P-FEC) and 2 were treated with paclitaxel monotherapy. C2P-RS was determined via C2P using frozen biopsy samples before NAC. RESULTS: Negative estrogen receptor (ER), negative progesterone receptor (PR), positive human epidermal growth factor receptor 2 (HER2), high Ki-67 expression and intermediate + high C2P-RS were significantly associated with high pathological complete response (pCR) rates compared with positive ER (30% versus 9%), positive PR (25% versus 6%), negative HER2 (34% versus 11%), low Ki-67 expression (24% versus 7%) or low C2P-RS (24% versus 9%), respectively. The combination of C2P-RS and Ki-67 had a stronger impact on pCR than each parameter alone, and a multivariate analysis showed that the combination was an independent predictor of pCR (odds ratio 3.3, 95% confidence interval 1.1-9.5). CONCLUSIONS: C2P-RS was significantly associated with pCR after P-FEC and may be a useful predictor for chemotherapy in BCs.

Clinicopathological analysis of GATA3-positive breast cancers with special reference to response to neoadjuvant chemotherapy.

BACKGROUND: The aim of this study was to investigate the clinicopathological characteristics of GATA binding protein 3 (GATA3)-positive breast cancers as well as the association of GATA3 expression with response to chemotherapy. PATIENTS AND METHODS: Tumor specimens obtained before neoadjuvant chemotherapy [paclitaxel followed by 5-fluorouracil/epirubicin/cyclophosphamide)] from breast cancer patients (n = 130) were subjected to immunohistochemical and mutational analysis of GATA3 and DNA microarray gene expression analysis for intrinsic subtyping. RESULTS: Seventy-four tumors (57%) were immunohistochemically positive for GATA3. GATA3-positive tumors were significantly more likely to be lobular cancer, estrogen receptor (ER)-positive, progesterone receptor (PgR)-positive, Ki67-negative, and luminal A tumors. Somatic mutations were found in only three tumors. Pathological complete response (pCR) was observed in 8 (11%) GATA3-positive tumors and in 22 (39%) GATA3-negative tumors. multivariate analysis showed that tumor size, human epidermal growth factor receptor 2 (her2), and gata3 were independent predictors of pcr. CONCLUSIONS: GATA3-positive breast cancers showed luminal differentiation characterized by high ER expression and were mostly classified as luminal-type tumors following intrinsic subtyping. Interestingly, GATA3 was an independent predictor of response to chemotherapy, suggesting that GATA3 might be clinically useful as a predictor of a poor response to chemotherapy.

Identification of essential sequence motifs in the node/notochord enhancer of Foxa2 (Hnf3beta) gene that are conserved across vertebrate species.

The expression of a winged-helix transcription factor, Foxa2/HNF3beta, is essential for development of the node and the notochord. We examined the node/notochord enhancer of mouse Foxa2 for sequence motifs conserved across vertebrate species. We cloned Foxa2 genes from chicken and fish, and identified the respective node/notochord enhancers that were active in transgenic mouse embryos. Comparison of the sequences of the enhancers revealed three evolutionally conserved sequence motifs, CS1, CS2 and CS3. Mutational analysis of the mouse enhancer indicated that CS3 is indispensable for gene expression in the node and the notochord, while CS1 and CS2 are required to augment enhancer activity. These motifs do not correspond to the consensus binding sequences of transcription factors known to be involved in node/notochord development.

Attenuating effect of a semipurified alcohol extract of soy protein on glomerular injury in spontaneous hypercholesterolemic male Imai rats.

We investigated whether soy protein's alcohol-extractable components (SPEs; mainly consisting of isoflavones) have the ability to attenuate glomerular injury in male Imai rats of a spontaneous focal segmental glomerulosclerosis model. Male Imai rats were fed a casein-based diet with and without SPEs. Group 1 (Cont) was fed a standard diet without additional SPEs, and groups 2 (SPE-1) and 3 (SPE-2) were fed a standard diet supplemented with a semipurified alcohol extract of soy protein, 0.05 and 0.10 g/100 g of diet, respectively. Body weight, urinary protein level, serum constituents, and systolic blood pressure were evaluated every 4 weeks from 12 through 28 weeks of age. At 28 weeks of age, rats were studied morphologically. Growth rates were not different among the three groups throughout the experiment. SPE-supplemented diets resulted in less proteinuria and less hyperlipidemia. The decline in renal function shown by blood urea nitrogen and creatinine clearance was less marked in the animals fed the SPE-supplemented diets. Each SPE-supplemented diet equally induced less glomerular hypertrophy and less renal histological damage compared with nonsupplemented diets. The present study showed a beneficial effect of a semipurified alcohol extract of soy protein on glomerular disease.

A specific thrombin inhibitor (argatroban) ameliorated cerebral blood flow in the patients with acute cerebral infarction.

Argatroban, one of the arginine derivatives, has been reported to have a safe and potent antithrombin action. This compound is active in several animal models of thrombosis and also has been shown to improve general neurological symptomatology, general subjective symptomatology and general daily behavior in the patients with acute thrombosis. This was considered to reflect remarkable improvement of microcirculation. No published clinical data, however, exist on the effect of argatroban on cerebral blood flow (CBF) change during acute stroke. Three patients with acute cerebral infarction were subjected to this study. Intravenous argatroban injection (2.5 mg/hr) was continued in 48 hours. Regional CBF (rCBF) was measured before and after injection of argatroban using Xe-CT method. Argatroban increased CBF not only in the injured side hemisphere or penumbra, but also contralateral side of lesion in the patients with acute cerebral infarction.

Bile alcohol profiles in bile, urine, and feces of a patient with cerebrotendinous xanthomatosis.

Bile alcohols in bile, urine, and feces of a patient with cerebrotendinous xanthomatosis have been analyzed by a combination of capillary gas-liquid chromatography and mass spectrometry after fractionation into groups according to mode of conjugation. The presence of at least 18 bile alcohols, which were excreted mainly as glucurono-conjugates in bile and urine, and as unconjugated forms in feces, was demonstrated. The following bile alcohols were identified with certainty by direct comparison with reference compounds: 5 beta-cholestane-3 alpha,7 alpha,12 alpha-triol; (23R)-5 beta-cholestane-3 alpha,7 alpha,12 alpha,23-tetrol; 5 alpha- and 5 beta-cholestane-3 alpha,7 alpha,12 alpha,24-tetrols; 5 alpha- and 5 beta-cholestane-3 alpha,7 alpha,12 alpha,25-tetrols; 27-nor-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24,25-pentol; (22R)-5 beta-cholestane-3 alpha,7 alpha,12 alpha,22,25-pentol; (23R)- and (23S)-5 beta-cholestane-3 alpha,7 alpha, 12 alpha,23,25-pentols; 3 alpha,12 alpha,25-trihydroxy-5 beta-cholestane-7-one; (24R)- and (24S)-5 beta-cholestane-3 alpha,7 alpha,12 alpha,24,25-pentols; 5 beta-cholestane-3 alpha,7 alpha,12 alpha,25,26-pentol. Although the bile alcohol profile in urine was quite different from those in bile and feces, the determination of urinary bile alcohols as well as of biliary and fecal bile alcohols could be used for diagnosis of cerebrotendinous xanthomatosis.

Modification of striatal arginine and citrulline metabolism by nitric oxide synthase inhibitors.

The effects of NG-substituted L-arginine (ARG) analogues on striatal ARG and citrulline (CIT) levels were investigated using in vivo microdialysis technique. A microdialysis probe was implanted into the striatum of anaesthesized Sprague-Dawley rats. Direct intrastriatal perfusion with 1 mM NG-nitro-L-arginine methyl ester (n = 8) increased striatal ARG release and decreased CIT release, suggesting suppressed NO synthase activity in the tissue. On the other hand, 1 mM NG-monomethyl-L-arginine (L-NMMA) (n = 6) evoked a persistent increase in both ARG and CIT. Considering that 4-320 microM L-ARG (n = 8) failed to increase CIT formation, CIT seems to be synthesized in the striatal tissue from L-NMMA by the enzyme that has been demonstrated in the kidney and aortic endothelium (NG,NG-dimethylarginine dimethyl-aminohydrolase).

Nitric oxide regulates NMDA-induced dopamine release in rat striatum.

We investigated the effect of nitric oxide (NO) on N-methyl-D-aspartate (NMDA)-induced changes in levels of dopamine (DA) and its metabolite in the striatum using in vivo microdialysis. Local administration of 1mM NMDA into the striatum significantly augmented DA release in the striatum. Simultaneous administration of -5mM N(G)-nitro-L-arginine methyl ester (-NAME), a NO synthase inhibitor, into the striatum significantly potentiated NMDA-induced DA release. This effect of L-NAME was completely reversed in the presence of 50mM L-arginine (L-Arg). Administration of 1 mM NMDA significantly decreased the levels of dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA). This effect of NMDA was not affected by concurrent administration of L-NAME. This study provides in vivo evidence for the involvement of NO in NMDA-induced DA release.

Presynaptic ionotropic glutamate receptors modulate in vivo release and metabolism of striatal dopamine, noradrenaline, and 5-hydroxytryptamine: involvement of both NMDA and AMPA/kainate subtypes.

In order to explore further the presynaptic modulation of monoamine release by glutamatergic nerve fibers, we investigated the effects of selective agonists for ionotropic glutamate (GLU) receptors on striatal release of dopamine (DA), noradrenaline (NA) and 5-hydroxytryptamine (5-HT). In the striatum of anesthetized Sprague-Dawley rats, in vivo microdialysis was performed to measure the release of monoamines and metabolities, and also to administer GLU agonists locally in the tissue. L-GLU and its selective agonists (N-methyl-D-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA) and kainate (KA)) evoked simultaneous release of striatal DA, NA and 5-HT in a dose-dependent manner. Pretreatment with MK-801 (5 mg/kg i.p.), a noncompetitive NMDA receptor antagonist, selectively suppressed NMDA-evoked monoamine release. The rank order of GLU agonist efficacy in releasing monoamines was different among DA, NA, and 5-HTergic terminals: AMPA = KA > NMDA for DA release, AMPA > NMDA = KA for NA release, and NMDA = AMPA = KA for 5-HT release. In conclusion, presynaptic ionotropic GLU receptors exist extensively on monoaminergic terminals including not only catecholaminergic (DA and NA) but also indoleaminergic (5-HT) terminals in the rat striatum. Their subtypes include both NMDA subtype and AMPA/KA subtype, and show a differential distribution among these three monoaminergic terminals and a differential contribution to facilitating monoamine release.

Correlation of in vivo nitric oxide and cGMP with glutamate/glutamine metabolism in the rat striatum.

We have examined how the suppression of endogenous production of nitric oxide (NO) in the striatal tissue affects release of glutamate (GLU) and glutamine (GLN) in pentobarbital-anesthetized male Sprague-Dawley rats. For the quantitative measurement of tissue NO production and amino acid release, an in vivo assay system for extracellular nitrite (NO2-) and amino acids was employed using an in vivo microdialysis technique. An NO synthase inhibitor (NG-nitro-L-arginine methyl ester, L-NAME) in concentrations ranging between 4-40 mM was perfused into the rat striatum using the assay system. Tissue NO production was found to be inversely proportional to the L-NAME concentration. L-NAME likewise decreased striatal levels of GLU and GLN. Furthermore, tissue NO production showed a positive correlation with GLU (R = 0.62, P < 0.02) and GLN (R = 0.86, P < 0.001) concentrations. Exogenous application of NO and cGMP by intrastriatal perfusion with 0.1-2.5 mM hydroxylamine and 0.4-10 mM 8-bromo-cGMP, respectively, increased striatal GLU release in a dose-related manner. Hydroxylamine reduced GLN release, and 8-bromo-cGMP showed a tendency to decrease GLN. In conclusion, striatal GLU/GLN metabolism is a function of the tissue concentration of NO. Normal endogenous concentration of NO causes GLU to be released at a consistent basal level, and enhanced tissue NO production facilitates GLU release via pathways including cGMP formation. We hypothesize that NO may suppress GLN formation by astrocytes.

Brain nitrite production during global ischemia and reperfusion: an in vivo microdialysis study.

Nitric oxide (NO) is considered to be associated with the pathogenesis of cerebral ischemic injury. In the present study, NO production was continuously monitored employing in vivo microdialysis. A microdialysis probe was inserted into the stratum. Levels of the major NO metabolite, NO-2, in the dialysate were determined using the Griess reaction. Rats were subjected to global cerebral ischemia produced by occlusion of both common carotid arteries together with induced hypotension. Cerebral ischemia induced a decrease in NO production, which was interrupted by a transient increase in NO synthesis. This increment was abolished in the presence of a NO synthase inhibitor, NG-nitro-L-arginine methyl ester (L-NAME), suggesting that NO synthase activity is transiently activated during ischemia. Following reperfusion, NO synthesis was enhanced. To our knowledge, this is the first report concerning the continuous temporal profile of NO production during global cerebral ischemia and reperfusion.

A novel in vivo assay system for consecutive measurement of brain nitric oxide production combined with the microdialysis technique.

A novel spectrophotometric nitrite (NO2-)/nitrate (NO3-) assay system for a small quantity (5 microliter) of dialysate sample obtained by in vivo brain microdialysis was developed based on the diazotization reaction. The system has the advantage of in vivo consecutive measurement, high precision, good reproducibility, technical simplicity, relatively short resolution time (2.5-20 min), and wide availability. The NO3- level in the rat striatum was found to be 3 times higher than the NO2- level. A nitric oxide (NO) synthase inhibitor, NG-nitro-L-arginine methyl ester, reduced striatal NO2-/citrulline formation in a dose-related manner and increased arginine, indicating that the tissue NO2- level detected by this assay system adequately reflects the striatal NO synthase activity.

Retinal ganglion cell response properties in the transcorneal electrically evoked response of the visual system.

To identify the retinal origin of a cortical evoked potential elicited by transcorneal electrical stimulation of the visual system (EER), the response properties of retinal ganglion cells (RGCs) of cats to transcorneal electrical stimuli were studied. The discharge latency of RGCs to transcorneal stimulation had two peaks with a high temporal resolution. The latency of early components of the EER is associated with the discharge latency of RGCs. Some RGCs showed prominent oscillatory discharges after transcorneal stimulation. Discharges of ON-bipolar cells responding to transcorneal stimulation were significantly inhibited by intravitreal injection of DL-2-amino-4-phosphonobutyrate (APB), which blocks the ON-pathway. These findings indicate that the EER has far-field potentials that might relate to oscillatory discharges of RGCs, and that ON bipolar cells and their related synaptic sites are involved in transcorneal electrical stimuli. The far-field potentials of the EER may have clinical applications, similar to those of somatosensoric evoked potentials and auditory brain stem potentials.

Bile acid profiles in bile, urine, and feces of a patient with cerebrotendinous xanthomatosis.

Bile acid profiles of bile, urine, and feces obtained from a patient with cerebrotendinous xanthomatosis on the same day have been analyzed by gas-liquid chromatography-mass spectrometry after fractionation into groups by mode of conjugation by an ion-exchange chromatography. The predominant biliary bile acid was cholic acid conjugated with glycine and taurine. Lesser amounts of the amino acid conjugates of chenodeoxycholic acid, ursodeoxycholic acid, 7-ketodeoxycholic acid, allocholic acid, and deoxycholic acid, and of unconjugated norcholic acid and allonorcholic acid were also present in the bile. The major fecal bile acid was 7-epicholic acid. Relatively large amounts of bile acids were excreted in the urine. Unconjugated 7-epicholic acid, norcholic acid, allonorcholic acid, and cholic acid predominated. The bile acid profiles of the patient were different from those of normal subjects and should be useful for the diagnosis.

Laparoscopic adrenalectomy: lateral transabdominal approach vs posterior retroperitoneal approach.

Laparoscopic adrenalectomy has been used to remove a wide variety of adrenal neoplasms. Although several laparoscopic approaches to the adrenal gland have been described, the lateral transabdominal approach has several advantages when compared with other approaches for laparoscopic adrenalectomy. From October 1995 to July 1999, we performed laparoscopic adrenalectomies on 16 patients, including eight posterior retroperitoneal approaches and eight lateral transabdominal approaches. Sixteen patients, ranging in age from 23 to 69 years, were treated for the following conditions: non-functioning adenoma, four patients; aldosteronoma, seven patients; pheochromocytoma, three patients; Cushing's adenoma, two patients. The average tumor size was 2.5 +/- 0.5 cm (1.8-3.0 cm, median 2.4 cm) in the lateral transabdominal approach, 1.2 +/- 0.8 cm (0.8-3.2 cm, median 1.75 cm) in the posterior retroperitoneal approach. Average operative time of lateral transabdominal approach was significantly shorter than that of the posterior retroperitoneal approaches (mean 129 min vs 269 min, P = 0.0005). Conversion to laparotomy was required in one patient in the posterior approach. Postoperative complication occurred in one pneumothorax in the lateral transabdominal approach and two subcutaneous emphysemas in the posterior retroperitoneal approach. There was no statistical difference in blood loss during the operation in the two groups. There was no mortality in either group. The lateral transabdominal approach is a safe and efficient technique for the removal of the adrenal neoplasms. Compared with other approaches, this technique has a wider working space and also good exposure for removing the adrenal gland.

Locus coeruleus stimulation exerts different influences on the dynamic changes of cerebral pial and intraparenchymal vessels.

The present experimental study was undertaken to investigate the effects of locus coeruleus stimulation on the dynamic changes of intraparenchymal vessels and pial vessels. Twelve cats were anaesthetized with alpha-chloralose and urethane. For stimulation of the locus coeruleus, a concentric stainless-steel needle electrode was inserted stereotaxically. During the stimulation, volumetric changes of the intraparenchymal vessels were monitored by a photoelectric method for estimating the cerebral blood volume (CBV) (6 cats), and the diameters of pial arteries were measured continuously using a video camera system (6 cats). The CBV followed a decreasing course during the stimulation of the locus coeruleus. The decrease in CBV from the control value (6.3 vol%) was 0.14 +/- 0.04 vol% at 80 s (p less than 0.05), 0.15 +/- 0.05 vol% at 100 s (p less than 0.05), and 0.15 +/- 0.03 vol% at 120 s (p less than 0.01). After cessation of the stimulation, CBV showed a gradual recovery. On the other hand, the diameters of the pial arteries did not change during or after the stimulation of the locus coeruleus. The above results suggest that the locus coeruleus has a vasoconstrictive effect on the intraparenchymal vessels, although it exerts no apparent influence on the pial arteries.

Anthophyllite exposure and endemic pleural plaques in Kumamoto, Japan.

OBJECTIVES: This study explored the high prevalence of pleural plaques in the town of Matsubase in Kumamoto, Japan. METHODS: Small-size chest X-ray film was used for screening, and all persons with pleural plaques were confirmed by computed tomography (CT). The prevalence rate of pleural plaques in the 4 districts of Matsubase and its surrounding towns and cities were also examined. The age-adjusted mortality rate for lung cancer in this town was compared with that of its surrounding towns and cities. RESULTS: Pleural plaques were found in 1357 persons (724 men and 633 women) among the inhabitants who were more than 20 years of age in Matsubase between 1988 and 1993. CT scans ascertained 938 cases with pleural plaques among the 11 14 persons who participated. Thus at least 9.5% of the inhabitants over 20 years of age in this town had pleural plaques. The neighboring towns had a higher rate than the more distant towns. A large-scale open-cast asbestos mine and mill had been in operation in Matsubase between 1883 and 1970. Mineral analysis revealed anthophyllite fibers. Most of the plaques were found in persons who had never worked in the mine or mill. CONCLUSIONS: The high prevalence of pleural plaques in Matsubase was due to anthophyllite exposure, mainly environmental. No mesotheliomas were found, however. These findings agree with those from an earlier study from Finland.

Ultrastructure, antigenicity, and histochemistry of stichocyte granules of adult Trichinella spiralis.

The stichosome of adult Trichinella spiralis was studied to determine its ultrastructural, antigenic, and histochemical characteristics. Stichocytes of adult worms had 2 types of granules, type I and type II, the ultrastructure of which was different from those of muscle larvae. Both types of granules consisted of a membrane surrounding a homogeneous matrix, and type I granules were rounder than type II granules. Sera from C3H mice immunized against excretory-secretory products of muscle larvae produced positive immunostaining of type I but not type II granules. Differences in antigenicity were observed between larval and adult stichocyte granules; monoclonal antibodies against alpha-granules of muscle larvae failed to label the adult granules. Azan staining revealed a histochemical difference between larval and adult stichocytes; adult stichocytes stained yellow, whereas larval stichocytes are known to stain red or blue. Thus, the present contribution revealed the existence of 2 distinct types of stichocyte granules in adult T. spiralis and showed them to differ profoundly from those characterized previously in muscle larvae.

A comparison of the results of A-C bypass grafting in collateral and non-collateral groups.

Ninety patients who had aorto-coronary bypass grafting were divided into two groups: a collateral group, which had coronary arterial stenosis or occlusion with collateral circulation, and a non-collateral group, which had coronary arterial stenosis or occlusion without collateral circulation. The number of coronary arteries visualized through collateral circulation in coronary angiograms (CAG) was 32, left anterior descending arteries (LAD) 17, right coronary arteries (RCA) 11, and left circumflex arteries (LCX) 4. The results of A-C bypass grafting in the collateral and non-collateral groups were compared. Surgical mortality was 0% in the collateral group, and 5.4% in the non-collateral group. The differences in graft patency and graft flow between the two groups were not statistically significant. However, left ventricular ejection fraction and myocardial perfusion, which was estimated by thallium-201 myocardial perfusion scintigram, were significantly improved after A-C bypass in the collateral group. Although the coronary arteries visualized through collateral vessels seemed too narrow to undergo graft anastomosis, they were, in fact, large enough. A-C bypass grafting was achieved with more satisfactory results in the collateral group than in the non-collateral group.