[Study on clinical phenotype of coal workers pneumoconiosis].

Objective: To explore the clinical phenotypic characteristics of coal worker's pneumoconiosis for guiding the individualized treatment of various types of patients with coal worker's pneumoconiosis. Methods: Collect clinical data of 121 cases of coal worker's pneumoconiosis in different stages, and select 16 clinical variables (age, smoking index, years of underground dust exposure, stages of pneumoconiosis, types of work, family history, main symptoms, secondary symptoms, CAT score, imaging manifestations, FVC%, FEV(1)/FVC, FEV(1)%, DLCO%, respiratory failure complications, pulmonary heart disease complications) . Principal Component Factor Analysis (PCA) was used to analyze 16 clinical variables of 121 patients with coal worker's pneumoconiosis. Extracted 2 principal components and 8 related variables from 16 clinical variables, then coal worker's pneumoconiosis patients were divided into three types according to CCC values. Variance analysis or χ(2) test were used to analyze the characteristics of these three types of clinical data, then summarized the clinical phenotype composition ratio and clinical data characteristics. Results: The patients with coal worker's pneumoconiosis were initially divided into three types, including 73 cases (60.3%) in type 1, 18 cases (14.9%) in type 2 and 30 cases (24.8%) in type 3. Patients in type 1 are mainly middle-aged, with little damage to lung function and mild clinical symptoms, the imaging manifestations of type 1 patients are mainly diffuse nodules, and the stages of pneumoconiosis are mostly one-stage and second-stage. Patients in type 2 are mainly in middle-aged and elderly patients.the main pulmonary impairment is diffuse function decline. The clinical symptoms are severe and the imaging manifestations are complex. The stages of pneumoconiosis are one, second and third stages. Patients in type 3 are mainly middle-aged and elderly patients, with more pulmonary function impairment (decreased ventilation and diffusion) , severe clinical symptoms, complex imaging manifestations (micro nodules, emphysema, mass shadow, fibrosis) , and those pneumoconiosis stages are mainly in the second and third stages. Conclusion: According to the clinical characteristics, the patients with coal worker's pneumoconiosis were divided into 3 types by cluster analysis method, the treatment plan has certain guiding value in clinical work according to different classifications.

[Clinical characteristics of pneumoconiosis complicated with chronic obstructive pulmonary disease].

Objective: To analyze the pulmonary function and clinical features of coal worker's pneumoconiosis complicated with chronic obstructive pulmonary disease (COPD) , coal worker's pneumoconiosis and COPD, in order to improve the diagnosis and treatment of coal worker's pneumoconiosis complicated with chronic obstructive pulmonary disease. Methods: Selected patients in respiratory department of General Hospital of Jincheng Coal Industry Group were classified as pneumoconiosis complicated with COPD group (n=52) , pneumoconiosis group (n=70) and COPD group (n=50) . Clinical data were collected and compared between three groups, including age, history of smoking, BMI, pulmonary function, CAT score and complication with Hypoxemia and respiratory faliure. Results: The mean age, smoking index and BMI of the three groups were not significantly different. The FEV1% pred, FEV(1)/FVC%, DLco-SB%, FVC% pred were significantly lower in pneumoconiosis complicated with COPD group than pneumoconiosis group (P<0.05) ; The FEV(1)% pred, DLco-SB%, FVC% pred were significantly lower in pneumoconiosis complicated with COPD group than COPD group (P<0.05) , but, the FEV(1)/FVC% was no significant different between pneumoconiosis complicated COPD group and COPD group (P>0.05) ; The CAT score for clinical symptoms of pneumoconiosis complicated with COPD group was significantly higher than that of pneumoconiosis group (P<0.05) , but there was no significant difference between pneumoconiosis complicated COPD group and COPD group (P>0.05) . The rate of hypoxemia in coal workers' pneumoconiosis combined with chronic obstructive pulmonary disease was 78.8%, which was higher than that of coal workers' pneumoconiosis group (61.4%) and chronic obstructive pulmonary disease group (72%) ; The respiratory failure rate of coal worker's pneumoconiosis combined with chronic obstructive pulmonary disease group was 44.2%, which was higher than that of coal worker's pneumoconiosis group (4.3%) and chronic obstructive pulmonary disease group (16%) . Conclusion: In pneumoconiosis patients, once complicate with COPD, the pulmonary function indexes are worse, the clinical symptoms are heavier, and the probability of hypoxemia and respiratory failure are higher. Compared with the COPD group, the patients with pneumoconiosis complicated with COPD have more restrictive ventilation dysfunction and diffuse dysfunction, and the clinical symptoms are heavier, and the probability of combined respiratory failure is higher.

Modulation of TLR2 and TLR4 in Macrophages Following Trichinella Spiralis Infection.

Parasitic helminthes can suppress and/or regulate the host immune response to allow long-term survival and chronic infection where toll-like receptors (TLRs) expressed on macrophages play essential roles in response to parasitic infection. Semi-quantitative PCR and flow cytometry studies about the modulation of TLRs and cytokine profiles in macrophages following T. spiralis infection were performed. TLRs, MyD88 and NF-κB were up-regulated by T. spiralis infection and essential to the parasite life cycles. Cytokines profiles (IL-6, IL-10, IL-12, TNF-α) were modulated during T. spiralis infection. Results suggest that T. spiralis infection may regulate the expression of TLR4 on macrophages and TLR4/MyD88/NF-κB signaling pathways. This study provides further insights into the mechanisms of TLR-mediated post-inflammatory response during T. spiralis infection.

Characterization of a new HLA allele: HLA-A*29:49.

The new A*29:49 differs from A*29:01:01:01 by one nucleotide at nt 368.

MiR-181 family: regulators of myeloid differentiation and acute myeloid leukemia as well as potential therapeutic targets.

MicroRNAs have been shown to play an important role in normal hematopoisis and leukemogenesis. Here, we report function and mechanisms of miR-181 family in myeloid differentiation and acute myeloid leukemia (AML). The aberrant overexpression of all the miR-181 family members (miR-181a/b/c/d) was detected in French-American-British M1, M2 and M3 subtypes of adult AML patients. By conducting gain- and loss-of-function experiments, we demonstrated that miR-181a inhibits granulocytic and macrophage-like differentiation of HL-60 cells and CD34+ hematopoietic stem/progenitor cells (HSPCs) by directly targeting and downregulating the expression of PRKCD (which then affected the PRKCD-P38-C/EBPα pathway), CTDSPL (which then affected the phosphorylation of retinoblastoma protein) and CAMKK1. The three genes were also demonstrated to be the targets of miR-181b, miR-181c and miR-181d, respectively. Significantly decreases in the expression levels of the target proteins were detected in AML patients. Inhibition of the expression of miR-181 family members owing to Lenti-miRZip-181a infection in bone marrow blasts of AML patients increased target protein expression levels and partially reversed myeloid differentiation blockage. In the mice implanted with AML CD34+ HSPCs, expression inhibition of the miR-181 family by Lenti-miRZip-181a injection improved myeloid differentiation, inhibited engraftment and infiltration of the leukemic CD34+ cells into the bone marrow and spleen, and released leukemic symptoms. In conclusion, our findings revealed new mechanism of miR-181 family in normal hematopoiesis and AML development, and suggested that expression inhibition of the miR-181 family could provide a new strategy for AML therapy.

Lack of involvement of neuronal nitric oxide synthase in the pathogenesis of a transgenic mouse model of familial amyotrophic lateral sclerosis.

A subset of familial cases of amyotrophic lateral sclerosis are linked to missense mutations in copper/zinc superoxide dismutase type 1. Patients with missense mutations in copper/zinc superoxide dismutase type 1 develop a paralytic disease indistinguishable from sporadic amyotrophic lateral sclerosis through an unknown toxic gain of function. Nitric oxide reacts with the superoxide anion to form the strong oxidant, peroxynitrite, which participates in neuronal injury in a variety of model systems. Peroxynitrite is an alternate substrate for copper/zinc superoxide dismutase type 1, causing catalytic nitration of tyrosine residues in other proteins. Mutations in copper/zinc superoxide dismutase type 1 may disrupt the active site of the enzyme and permit greater access of peroxynitrite to copper, leading to increased nitration by peroxynitrite of critical cellular targets. To investigate whether neuronal-derived nitric oxide plays a role in the pathogenesis of familial amyotrophic lateral sclerosis, we examined the effects of three different nitric oxide synthase inhibitors: a non-selective nitric oxide synthase inhibitor, nitro-L-arginine methyl ester; a relatively selective inhibitor of neuronal nitric oxide synthase, 7-nitroindazole; and a novel highly selective neuronal nitric oxide synthase inhibitor, AR-R 17,477, in transgenic mice expressing a familial amyotrophic lateral sclerosis-linked mutant human copper/zinc superoxide dismutase type 1 (Gly-->Ala at position 93; G93A) containing a high transgene copy number and a low transgene copy number. AR-R 17,477, but not nitro-L-arginine methyl ester or 7-nitroindazole, significantly prolonged survival in both the high and low transgene transgenic mice. To determine whether neuronal nitric oxide synthase is involved in the pathogenesis resulting from the familial amyotrophic lateral sclerosis copper/zinc superoxide dismutase type 1 mutation, we produced mice with the copper/zinc superoxide dismutase type 1 mutation which lack the neuronal nitric oxide synthase gene. The transgenic mice expressing a familial amyotrophic lateral sclerosis-linked mutant human copper/zinc superoxide dismutase type 1 on neuronal nitric oxide synthase null background do not live significantly longer than transgenic mice expressing a familial amyotrophic lateral sclerosis-linked mutant human copper/zinc superoxide dismutase type 1. Western blot analysis indicates the presence of two neuronal nitric oxide synthase-like immunoreactive bands in spinal cord homogenates of the neuronal nitric oxide synthase null mice, and residual neuronal nitric oxide synthase catalytic activity ( > 7%) is detected in the spinal cord of the transgenic mice expressing a familial amyotrophic lateral sclerosis-linked mutant human copper/zinc superoxide dismutase type 1 on neuronal nitric oxide synthase null background. This amount of residual activity probably does not account for lack of protection afforded by the disrupted neuronal nitric oxide synthase gene in the familial amyotrophic lateral sclerosis-linked mutant human copper/zinc superoxide dismutase type 1 mice. Immunological nitric oxide synthase is not detected in the copper/zinc superoxide dismutase type 1 mutant mice at several different ages, thus excluding immunological nitric oxide synthase as a contributor to the pathogenesis of familial amyotrophic lateral sclerosis. Levels of neuronal nitric oxide synthase as well as Ca2+-dependent nitric oxide synthase catalytic activity in the copper/zinc superoxide dismutase type 1 mutant mice do not differ from wild type mice. Endothelial nitric oxide synthase levels may be decreased in the copper/zinc superoxide dismutase type 1 mutant mice. Together, these results do not support a significant role for neuronal-derived nitric oxide in the pathogenesis of familial amyotrophic lateral sclerosis transgenic mice.

Recombinant adenovirus is an appropriate vector for endocytotic protein trafficking studies in cultured neurons.

Endocytosis of full-length beta-amyloid precursor protein (APP) from the plasma membrane contributes to beta-amyloid peptide (Abeta) secretion, and, hence, potentially contributes to the molecular pathogenesis of Alzheimer's disease. We recently have demonstrated that central neuronal APP is endocytosed in a common vesicular compartment with recycling synaptic vesicle integral membrane proteins, but is then sorted away from synaptic vesicles for retrograde transport to the neuronal soma. For this report, we explore whether recombinant adenovirus can be used to modulate APP expression in cultured central neurons to study APP processing by the endocytotic pathway in these cells. Using a replication-deficient recombinant adenovirus that expresses a lacZ reporter (Ad5/CMV-lacZ), we demonstrate high efficiency of transfection (30-35%) at low viral titer (10-20 MOI), with no significant neuronal toxicity or cytoarchitectural change. In addition, we demonstrate that infection with the control virus does not result in re-direction of endogenous neuronal APP from usual endocytotic pathways. We have prepared, using the same genomic background as the control virus, an adenoviral vector that expresses the neuronal isoform of human APP (Ad5/CMV-APP). Infection with Ad5/CMV-APP at 10-20 MOI results in significantly increased immunoreactivity for endocytosed APP with preservation of usual endocytotic trafficking. These results demonstrate that recombinant adenovirus at low titer is an appropriate and effective vector for protein trafficking/processing studies in cultured central neurons.

Thrombogenicity of polyethylene oxide-bonded Dacron sewing ring in a mechanical heart valve.

BACKGROUND AND AIM OF THE STUDY: The aim of the study was to evaluate the effect of binding hydrophilic polyethylene oxide (PEO) onto Dacron fibers in the sewing ring of a mechanical heart valve (MHV), in terms of thrombogenicity of the prosthesis. METHODS: The study was performed in blinded fashion. Six Yorkshire-cross pigs (bodyweight 35-45 kg) were implanted with MHVs, in the mitral annulus, with the PEO-treated sewing ring. An additional five pigs implanted with identical MHVs, but with untreated sewing rings, served as controls. PEO of chain-length 10,000 Da was grafted to Dacron fibers using gamma irradiation. PEO-bonded Dacron fibers (diameter 100 microns) were used to weave the sewing ring, which was then assembled on a titanium stent (OD 25 mm). Autologous platelets were labeled with 111In-tropolone and injected intravenously (850-1250 microCi per injection) into the pigs on removal from cardiopulmonary bypass (CPB). At 20-24 h after surgery, platelet thrombi adherent to MHV components, and shed emboli trapped in the brain, lung, heart, kidneys and other organs/connective tissues were imaged using a gamma camera. The animals were killed and the amounts of thrombi adherent to MHV components and organ-trapped emboli quantified using an ionization chamber and gamma counter. RESULTS: There was no statistically significant difference in the adhesion of 111In-labeled platelets to either control sewing rings (0.08 +/- 0.06% dose) or PEO-treated rings (0.19 +/- 0.21% dose). The thrombogenicity of MHV components in both animal groups was in the ascending order: Dacron ring > Teflon pledgets > polypropylene sutures > titanium housing > pyrolytic carbon. The number of platelet-emboli trapped in the organs was not significantly different between the two groups. CONCLUSIONS: Simple modifications may not reduce platelet thrombosis or wound-healing of the sewing ring in the acute phase, at which time several complex processes are activating and inactivating platelets and coagulant factors during CPB and implantation of MHVs.

A new method for quantitation of platelet microthrombi and microemboli from cardiopulmonary bypass in organs using 111In labeled platelets.

During cardiopulmonary bypass (CPB), showers of microemboli (ME) distribute among the organs and connective tissues according to regional blood flow. Post CPB, ME were quantified by subtracting residual platelets (RP) in the organs of a group of unoperated control Yorkshire pigs (n = 6) from those of operated pigs. The RP level was minimized by heparinization (300 IU/kg) before death and exsanguination. The number of adherent microthrombi (MT) and ME from the oxygenator (OX), arterial filter (AF), and thoracotomy site were determined using 111In labeled autologous platelets (INPLT) (525-585 microCi administered 24 hr before CPB) in two CPB groups (ACT > 400 sec) of 12 pigs (30-35 kg). CPB was carried out at a flow of 2.5-3.5 L/min at 28 degrees C with a roller or a centrifugal pump, OX (Bentley Univox 1.8 m2), AF (0.25 m2), and cardiotomy reservoir (CR) (Bentley BR: 3,500), for 90 (n = 6) and 180 (CPB 180, n = 6) min. Six pigs underwent thoracotomy without CPB. L-Arginine was infused at a dose of 2 mg/ kg/min during CPB (n = 6). Flow cytometry was used to estimate the circulating ME in blood. MT and organ trapped ME were imaged with a gamma camera and measured with an ion chamber and a gamma counter. ME values (percent of injected INPLT dose) in six organs and four connective tissues were calculated for all five groups. INPLT distribution indicated a uniform distribution of low level platelet MT in the CR and AF. Circulating ME amounted to 2.5% of total platelets. In the CPB circuit, ME generation in AF was the rate-limiting step (n = 4 x 10(5)). Similar studies in organs and tissues suggested the presence of a uniform distribution of the total events of ME (n = 500 x 10(6)). ME increase in brain, lung, liver, and skeletal muscle following thoracotomy and CPB was significant. The low level of ME in ischemia sensitive organs also indicated the presence of a thrombolytic threshold for cumulative ME. ME disaggregation was activated at an early stage to prevent ischemic damage, specifically in the brain. Measurement of trapped ME provided a novel, reliable, and one step method of evaluation of thrombogenicity of a CPB device and drugs.

Successful prosthetic mitral valve implantation in pigs.

Clotting mechanisms, the coagulation cascade, platelet function, and platelet-leukocyte-endothelial cell interactions are all very similar in humans and pigs. Because of these similarities, the authors concluded that the pig would be an ideal model for the study of thromboembolism resulting from prosthetic heart valves. To date, they have successfully recovered a total of 11 pigs (52.9 +/- 8.1 kg), 3 with bioprosthetic valves and 8 with mechanical valves, all in the mitral position (25 mm od). The normal presence of high numbers of pulmonary endothelial macrophages and other unique aspects of porcine cardiovascular and pulmonary function dictate somewhat different surgical protocols than those normally used for human patients and ruminant species. Some of these special procedures include 1) crystalloid prime without the use of plasma volume expanders, especially those with a starch base; 2) pharmacologic protection against arrhythmias (lidocaine, 4 mg/kg); 3) special attention to adequate hypothermic cardioprotection during the time of cross-clamp; 4) the use of shock doses of corticosteroid (prednisolone sodium succinate, 0.5 mg/kg) before removal of the aortic cross-clamp; and 5) positive inotropic support (dopamine, 0.008 mg/kg) while weaning from cardiopulmonary bypass. Gamma camera images of 111In tagged autologous platelets 24 hours after surgery show most thrombi located on the sewing ring with fewer on the pledgets and anchor sutures. The latter observations were confirmed by quantification of platelet deposition using a gamma counter.

Enhancer, silencer, and growth factor responsive regulatory sequences in the promoter for the mouse choline acetyltransferase gene.

We describe here the isolation of the 5' region of the mouse choline acetyltransferase (ChAT) gene and the functional characterization of regulatory regions that control its expression. ChAT catalyzes the synthesis of a neurotransmitter, acetylcholine, and is expressed specifically in cholinergic neurons. The 5' flanking region of the mouse ChAT gene lacks a consensus TATA element and transcription initiates at multiple sites. The gene contains a strong enhancer in the promoter-proximal region, a weaker, nerve growth factor responsive enhancer located immediately upstream, and a silencer-like sequence yet further upstream. We identified the strong enhancer within the proximal promoter region -430 to -115 by its ability to activate a heterologous, minimal SV40 promoter. The ChAT enhancer functioned in both a position- and orientation-independent manner. The region from -825 to -430 contained weaker enhancer activity, but was modulated more strongly by exposure of transfected cells to NGF. The activity of the strong, -430 to -115 enhancer was not modulated by NGF. In our experiments, NGF coordinately increased expression of endogenous ChAT enzymatic activity and expression of reporter genes placed under the control of the ChAT promoter and enhancers. Activity of the ChAT proximal promoter is silenced in noncholinergic neuronal cell lines (B103 and F11) and in nonneuronal L6 myoblasts by an upstream region from -925 to -558. These results indicate that regulation of the ChAT gene is achieved by a combination of both positive and negative regulatory mechanisms whose interaction determines the expression of a cholinergic neuronal phenotype.

Pathogenic mechanisms in familial amyotrophic lateral sclerosis due to mutation of Cu, Zn superoxide dismutase.

Oxidative mechanisms of damage have been implicated indirectly in the damage to brain tissue caused acutely by ischemia or chronically by neurodegenerative diseases. A direct link between pathogenesis and antioxidant enzyme systems has come from studies of a genetic form of amyotrophic lateral sclerosis (ALS). ALS causes the degeneration of motor neurons in cortex, brainstem and spinal cord with consequent progressive paralysis and death. The disease occurs in both sporadic and familial forms. Some 20% of kindreds in which ALS is inherited in an autosomal dominant fashion have mutations in the gene (SOD1) encoding Cu, Zn superoxide dismutase (SOD). Several SOD1 mutations have been shown by ourselves and others to cause motor neuron disease when expressed at high levels in transgenic mice, whereas transgenic mice expressing comparable amounts of wild-type human SOD do not show clinical disease. Thus, we have argued that motor neuron disease is caused by gain-of-function mutations in the human SOD1 gene. Our current experiments investigate the link between mutation of SOD1 and oxidative pathways of damage.

Effects of deletion of streptokinase residues 48-59 on plasminogen activation.

Streptokinase (SK) is a thrombolytic agent widely used for the clinical treatment of clotting disorders such as heart attack. The treatment is based on the ability of SK to bind plasminogen (Pg) or plasmin (Pm), forming complexes that proteolytically activate other Pg molecules to Pm, which carries out fibrinolysis. SK contains three major domains. The N-terminal domain, SKalpha, provides the complex with substrate recognition towards Pg. SKalpha contains a unique mobile loop, residues 45-70, absent in the corresponding domains of other bacterial Pg activators. To study the roles of this loop, we deleted 12 residues in this loop in both full-length SK and the SKalpha fragment. Kinetic data indicate that this loop participates in the recognition of substrate Pg, but does not function in the active site formation in the activator complex. Two crystal structures of the deletion mutant of SKalpha (SKalpha(delta)) complexed with the protease domain of Pg were determined. While the structure of SKalpha(delta) is essentially the same as this domain in full-length SK, the mode of SK-Pg interaction was however different from a previously observed structure. Even though mutagenesis studies indicated that the current complex represents a minor interacting form in solution, the binding to SKalpha(delta) triggered similar conformational changes in the Pg active site in both crystal forms.

Primary lateral sclerosis: A heterogeneous disorder composed of different subtypes?

OBJECTIVE: To determine identifiable subgroups of patients with primary lateral sclerosis (PLS) with distinct clinical features as a first step in identifying patients likely to have the same disorder. METHODS: Twenty-five patients meeting previously proposed diagnostic criteria for PLS were seen for examination, measurement of gait and finger tapping speed, and physiologic tests to assess motor pathways. Motor cortex excitability and central motor conduction time were assessed with transcranial magnetic stimulation. Brainstem motor pathways were assessed by the acoustic startle reflex. MRS was performed in a subgroup of patients to assess metabolites in the motor cortex. RESULTS: Fifty-six percent of the patients with PLS had a similar pattern of symptom progression, which the authors termed ascending. In these patients spasticity began in the legs and progressed slowly and steadily. Spasticity in the arms developed 3.6 years after the legs, on average, and speech impairment followed 1.5 years later. Motor evoked potentials were absent. MRS showed a mean reduction of N-acetylaspartate/creatinine in the motor cortex. The remaining patients with PLS had heterogeneous patterns of symptom progression and physiology. CONCLUSIONS: Patients with PLS with an ascending progression of symptoms form a distinct clinical subgroup that may be amenable to investigations of etiology and treatment.

Myocardial ischemia-reperfusion injury in estrogen receptor-alpha knockout and wild-type mice.

We investigated the function of estrogen receptor-alpha in global myocardial ischemia and reperfusion injury in male estrogen receptor-alpha knockout (ERKO) and wild-type mice. Mouse hearts were subjected to 45 min of global ischemia followed by 180 min of reperfusion. The hearts were excised, cannulated, and maintained in a chilled (4 degrees C) cardioplegia solution until warm (37 degrees C) oxygenated Krebs-Henseleit bicarbonate buffer was perfused through the coronary arteries. ERKO hearts started beating later and had a higher incidence of ventricular fibrillation and/or tachycardia than control hearts. Coronary flow rate was significantly lower in ERKO hearts during the 90- and 120-min periods of reperfusion. Ca(2+) accumulation was significantly greater following 30, 90, 120, 150, and 180 min of reperfusion in ERKO hearts. Nitrite production was significantly less in ERKO hearts following 90, 120, and 150 min of reperfusion. Myocardial reduction of 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide was significantly lower in experimental ERKO hearts. Marked interstitial edema and contraction bands were seen in hematoxylin-eosin-stained sections of ischemia-reperfused ERKO hearts but not in control tissues. Hematoxylin-basic fuchsin-picric acid-stained sections from experimental ERKO hearts had fewer viable myocytes compared with controls. Transmission electron microscopy revealed swollen and fragmented mitochondria with amorphous and granular bodies, loss of matrix, and rupture of cristae in experimental ERKO hearts. This is the first demonstration that estrogen receptor-alpha plays a cardioprotective role in ischemia-reperfusion injury in males.

Effects of dietary phytoestrogen on global myocardial ischemia-reperfusion injury in isolated female rat hearts.

We investigated the effects of phytoestrogen on global myocardial ischemia-reperfusion injury in five groups of female rats. A high-phytoestrogen group (HPE) was ovariectomized (Ovx) and fed a diet containing soybean protein and a high-isoflavone soy extract. Another Ovx group of rats was fed the same diet as the HPE group but treated with the estrogen receptor blocker ICI-182,780 (HPE + ICI). A third group of Ovx rats was fed a diet containing soybean protein alone (low-phytoestrogen content; LPE). A fourth Ovx group was fed a diet free of phytoestrogen (Ovx). The fifth group of rats was sham ovariectomized (sham). Hearts from all rats were subjected to 30 min of global, hypothermic (4 degrees C), cardioplegic ischemia and 120 min of normothermic (37 degrees C) reperfusion with oxygenated Krebs-Henseleit buffer. Compared with either the sham or the HPE group, the Ovx and HPE + ICI groups had significantly decreased first derivative of left ventricular pressure (dP/dt), coronary flow rate (CFR), nitrite production and mitochondrial respiratory function and significantly increased Ca2+ accumulation and myocardial histological and ultrastructural injury. The CFR of the LPE group was significantly different from that of either Ovx or HPE + ICI group but the dP/dt, nitrite production, Ca2+ accumulation, and mitochondrial function were not. Our results indicate that diets containing phytoestrogen extract play a cardioprotective role in global myocardial ischemia-reperfusion in female rats.

Effect of estrogen on global myocardial ischemia-reperfusion injury in female rats.

We investigated the effects of estrogen on global myocardial ischemia-reperfusion injury in rats that were ovariectomized (Ovx), sham-operated, or ovariectomized and then given 17beta-estradiol (E(2)beta) supplementation (Ovx+E(2)beta). Hearts were excised, cannulated, perfused with and then immersed in chilled (4 degrees C) cardioplegia solution for 30 min, and then retrogradely perfused with warm (37 degrees C), oxygenated Krebs-Henseleit bicarbonate buffer for 120 min. The coronary flow rate, first derivative of left ventricular pressure, and nitrite production were all significantly lower in Ovx than in sham-operated or Ovx+E(2)beta hearts. However, coronary flow rates or nitrate production were not consistently different throughout the entire reperfusion period. Ca(2+) accumulated more in Ovx rat hearts than in sham-operated or Ovx+E(2)beta hearts, and mitochondrial respiratory function was lower in Ovx hearts than in hearts from the other two groups. Marked interstitial edema and contraction bands were seen in hematoxylin-eosin-stained sections of Ovx rat hearts but not in hearts from either of the other groups. Hematoxylin-basic fuchsin-picric acid-stained sections revealed fewer viable myocytes in hearts from the Ovx group than from the sham or Ovx+E(2)beta group. Transmission electron microscopy demonstrated more severely damaged mitochondria and ultrastructural damage to myocytes in Ovx rat hearts. Our results indicate that estrogen plays a cardioprotective role in global myocardial ischemia-reperfusion injury in female rats.

Benefit of vitamin E, riluzole, and gabapentin in a transgenic model of familial amyotrophic lateral sclerosis.

Familial amyotrophic lateral sclerosis (FALS) has been linked in some families to dominant mutations of the SOD1 gene encoding Cu,Zn superoxide dismutase (Cu,ZnSOD). We have used a transgenic model of FALS based on expression of mutant human Cu,ZnSOD to explore the etiology and therapy of the genetic disease. Expression of mutant, but not wild-type, human Cu,ZnSOD in mice places the brain and spinal cord under oxidative stress. This causes depletion of vitamin E, rather than the typical age-dependent increase in vitamin E content as occurs in nontransgenic mice and in mice expressing wild-type human Cu,ZnSOD. Dietary supplementation with vitamin E delays onset of clinical disease and slows progression in the transgenic model but does not prolong survival. In contrast, two putative inhibitors of the glutamatergic system, riluzole and gabapentin, prolong survival. However, riluzole did not delay disease onset. Thus, there was clear separation of effects on onset, progression, and survival by the three therapeutics tested. This suggests the hypothesis that oxidative damage produced by the expression of mutant Cu,ZnSOD causes slow or weak excitotoxicity that can be inhibited in part by alerting glutamate release or biosynthesis presynaptically.

Age-dependent penetrance of disease in a transgenic mouse model of familial amyotrophic lateral sclerosis.

The mutation gly93-->ala of Cu,Zn superoxide dismutase (SOD) is found in patients with familial amyotrophic lateral sclerosis and causes motor neuron disease when expressed in transgenic mice. The progression of clinical and pathological disease was studied in a line of mice designated G1H. Clinical disease started at 91 +/- 14 days of age with fine shaking of the limbs, followed by paralysis and death by 136 +/- 7 days of age. Pathological changes begin by 37 days of age with vacuoles derived from swollen mitochondria accumulating in motor neurons. At the onset of clinical disease (90 days), significant death of somatic motor neurons innervating limb muscles has occurred; mice at end-stage disease (136 days) show up to 50% loss of cervical and lumbar motor neurons. However, neither thoracic nor cranial motor neurons show appreciable loss despite vacuolar changes. Autonomic motor neurons also are not affected. Mice that express wild-type human Cu,Zn SOD remain free of disease, indicating that mutations cause neuron loss by a gain-of-function. Thus, the age-dependent penetrance of motor neuron disease in this transgenic model is due to the gradual accumulation of pathological damage in select populations of cholinergic neurons.