Stretching DNA with optical tweezers

Force-extension (F-x) relationships were measured for single molecules of DNA under a variety of buffer conditions, using an optical trapping interferometer modified to incorporate feedback control. One end of a single DNA molecule was fixed to a coverglass surface by means of a stalled RNA polymerase complex. The other end was linked to a microscopic bead, which was captured and held in an optical trap. The DNA was subsequently stretched by moving the coverglass with respect to the trap using a piezo-driven stage, while the position of the bead was recorded at nanometer-scale resolution. An electronic feedback circuit was activated to prevent bead movement beyond a preset clamping point by modulating the light intensity, altering the trap stiffness dynamically. This arrangement permits rapid determination of the F-x relationship for individual DNA molecules as short as -1 micron with unprecedented accuracy, subjected to both low (approximately 0.1 pN) and high (approximately 50 pN) loads: complete data sets are acquired in under a minute. Experimental F-x relationships were fit over much of their range by entropic elasticity theories based on worm-like chain models. Fits yielded a persistence length, Lp, of approximately 47 nm in a buffer containing 10 mM Na1. Multivalent cations, such as Mg2+ or spermidine 3+, reduced Lp to approximately 40 nm. Although multivalent ions shield most of the negative charges on the DNA backbone, they did not further reduce Lp significantly, suggesting that the intrinsic persistence length remains close to 40 nm. An elasticity theory incorporating both enthalpic and entropic contributions to stiffness fit the experimental results extremely well throughout the full range of extensions and returned an elastic modulus of approximately 1100 pN.     

T-strand integration in maize protoplasts after codelivery of a T-DNA substrate and virulence genes

We describe a plant protoplast transformation method that provides transformants with a simple pattern of integration of a foreign gene. The approach is to deliver into plant protoplasts by direct gene transfer the Agrobacterium virulence genes virD1 and virD2 with or without virE2, together with a target plasmid containing a gene of interest flanked by Agrobacterium T-DNA border repeat sequences of 25 bp. We present evidence of T-DNA formation in maize protoplasts and its integration into the maize genome. The frequency of VirD1-VirD2-mediated integration events was about 20-35% of the total number of transformants. The addition of virE2 doubled the transformation efficiency. The method described here is of sufficient efficiency and simplicity to be useful for the production of transgenic plants with single-copy well-defined transgenic inserts.     

Electroconvulsive therapy for treatment of intractable seizures. Initial findings in two children

We treated two children with intractable epilepsy with electroconvulsive therapy (ECT) for seizure control. One child showed a change in seizure pattern with treatment, which at greater intensity was also effective in stopping nonconvulsive status epilepticus. The other child showed a decrease in spontaneous seizure frequency during short-term treatment. These findings suggest a possible role for ECT in the management of intractable epilepsy in children who are not candidates for epileptic surgery.     

A novel approach for comparative study of periosteum, muscle, subcutis, and skin microcirculation by intravital fluorescence microscopy

Herein, we report a new model, which allows comparative study of the microcirculation of different peripheral tissues, i.e., periosteum, skeletal muscle, subcutis, and skin. Using dextran-insensitive Wistar rats gracilis and semitendinosus muscles of the left hindlimb were prepared in association with their appertaining tibial fragments, subcutis, and skin. Blood supply was guaranteed by the femoral artery via the saphenous vessels. High-resolution intravital epi-illumination microscopy of the two muscles displayed the typical microvascular architecture with the capillaries running in parallel to each other (capillary density (CD) 128.4 +/- 4.5 cm-1). In subcutis and skin, capillaries were found arranged as interconnecting mesh-like networks with a density, which was significantly higher (P < 0.05) in subcutis (191.0 +/- 5.5 cm-1) compared with skin (108.9 +/- 3.3 cm-1). Analysis of periosteal tissue revealed two distinct types of arrangements of microvascular architecture. Adjacent to the major feeding and draining vessels of the periosteum, capillaries were organized in densely meshed shunt-like networks, revealing the highest capillary density (242.7 +/- 13.2 cm-1; P < 0.05) of all tissues studied. Periosteal capillaries distant from the major feeding and draining vessels were arranged in parallel to the longitudinal axis of the tibial bone and presented with a density similar to that of the skeletal muscle (128. 6 +/- 9.4 cm-1). Topical application of acetylcholine for analysis of physiological reactivity of the microvasculature showed dose-dependent arteriolar dilation. Moreover, a 3-min upstream femoral artery occlusion demonstrated an appropriate hyperemic response in all tissues studied, indicating intact myogenic control. A prolonged period of ischemia (120 min) followed by reperfusion (60 min) caused massive (P < 0.05) leukocyte-endothelial cell interaction in postcapillary venules, similarly as reported in other microvascular tissue preparations. We propose that the model presented provides a good approach to all peripheral tissues for both the analysis of the physiology of tissue-confined microvascular control and the development of novel therapeutic strategies to counteract manifestation of nutritional dysfunction and inflammatory response in disease.     

Phase II study of receptor-enhanced chemosensitivity using recombinant humanized anti-p185HER2/neu monoclonal antibody plus cisplatin in patients with HER2/neu-overexpressing metastatic breast cancer refractory to chemotherapy treatment

PURPOSE: To determine the toxicity, pharmacokinetics, response rate, and response duration of intravenous (i.v.) administration of recombinant, humanized anti-p185HER2 monoclonal antibody (rhuMAb HER2) plus cisplatin (CDDP) in a phase II, open-label, multicenter clinical trial for patients with HER2/neu-overexpressing metastatic breast cancer. PATIENTS AND METHODS: The study population consisted of extensively pretreated advanced breast cancer patients with HER2/neu overexpression and disease progression during standard chemotherapy. Patients received a loading dose of rhuMAb HER2 (250 mg i.v.) on day 0, followed by weekly doses of 100 mg i.v. for 9 weeks. Patients received CDDP (75 mg/m2) on days 1, 29, and 57. RESULTS: Of 37 patients assessable for response, nine (24.3%) achieved a PR, nine (24.3%) had a minor response or stable disease, and disease progression occurred in 19 (51.3%). The median response duration was 5.3 months (range, 1.6-18). Grade III or IV toxicity was observed in 22 of 39 patients (56%). The toxicity profile reflected that expected from CDDP alone with the most common toxicities being cytopenias (n = 10), nausea/vomiting (n = 9), and asthenia (n = 5). Mean pharmacokinetic parameters of rhuMAb HER2 were unaltered by coadministration of CDDP. CONCLUSION: The use of rhuMAb HER2 in combination with CDDP in patients with HER2/neu-overexpressing metastatic breast cancer results in objective clinical response rates higher than those reported previously for CDDP alone, or rhuMAb HER2 alone. In addition, the combination results in no apparent increase in toxicity. Finally, the pharmacology of rhuMAb HER2 was unaffected by coadministration with CDDP.     

Negative predictive value of imaging-guided abdominal biopsy results: cytologic classification and implications for patient management

OBJECTIVE: Our purpose was to assess the negative predictive value of imaging-guided abdominal biopsy results and correlate it with cytology classification, lesion size, needle gauge, and cancer history. MATERIALS AND METHODS: A retrospective study was performed of 100 patients with proven diagnoses who had undergone imaging-guided abdominal biopsies showing no cells that were malignant or suspicious for malignancy. Specimens were classified as normal or benign, nondiagnostic, or atypical. Negative predictive value was calculated for each cytologic category, lesion size, needle gauge, and cancer history. Logistic regression analysis was performed to allow us to identify predictors of false-negative results. RESULTS: Overall negative predictive value was 67%. Other negative predictive values were normal or benign result, 78%; nondiagnostic result, 66%; and atypical result, 29%. Negative predictive value was greater when the lesion was large (> or =3 cm) (p = .031). Logistic regression analysis allowed us to predict a 9.3% chance of a false-negative result for a specimen of normal or benign cytology that was taken from a large lesion in a patient with no cancer history but an 87% chance of a false-negative when a specimen of atypical cytology was taken from a small (<3 cm) lesion in a patient with a cancer history. CONCLUSION: Imaging-guided abdominal biopsy specimens containing atypical cells should be viewed with caution. In patients without cancer, if a lesion is large and the specimen contains normal target organ or benign cells, the likelihood of a false-negative result may be low enough that imaging surveillance at appropriate intervals may be sufficient.     

An agency-based representative payee program and improved community tenure of persons with mental illness

OBJECTIVE: Representative payee programs help severely mentally ill individuals manage money from their Social Security payments to cover expenses for necessities and to avoid homelessness and rehospitalization. This study examined a representative payee program operated by a community mental health center to determine the criteria used by clinicians and ease managers to refer clients to the program and to learn whether participation in the program was associated with reductions in hospitalization. METHODS: The retrospective study included 56 individuals with severe mental illness who were enrolled in the representative payee program at Community Counseling Centers of Chicago for one year and who also had received services from the agency for at least one year before enrollment. Criteria used to refer clients to the representative payee program were determined through chart reviews. Data on state hospitalizations before and after enrollment were available for the entire sample; additional data on Medicaid-funded private hospitalizations were available for a subset of 33 clients. RESULTS: The most common criteria for enrollment in the representative payee program were comorbid substance abuse or dependence (49 percent), a history of homelessness (33 percent), and frequent hospitalizations (32 percent). During the year of participation in the representative payee program, the mean number of days spent in state hospitals decreased markedly compared with the year before enrollment, from 68 days to seven days. A similar reduction was noted in the number of days spent in state and private hospitals, from 97 days to 15 days. CONCLUSIONS: Findings from this pre- and postintervention retrospective study are tentative in the absence of a more rigorous design. However, the results suggest that the representative payee program is quite effective in reducing hospital stays.     

Dietary monounsaturated fatty acids promote aortic atherosclerosis in LDL receptor-null, human ApoB100-overexpressing transgenic mice

In mice with genetically engineered high levels of plasma low density lipoprotein (LDL), we tested the hypothesis that an increase in the dietary content of monounsaturated fatty acids but not of polyunsaturated fatty acids would promote atherosclerosis. The mouse model used was an LDL receptor-null, human apoB100-overexpressing strain. Six experimental groups of 19 to 38 mice of both sexes were established when the animals had reached 8 weeks of age. For the next 16 weeks, individual groups were fed either a commercial diet or prepared diets including fat as 10% of energy, with 5 different fatty acid enrichment patterns including the following: saturated (sat), cis and trans monounsaturated (mono), and n-3 and n-6 polyunsaturated (poly). Highly significant differences (ANOVA, P<0. 0001) in LDL cholesterol (in mg/dL) were found, with the rank order at 16 weeks being trans mono (mean, 1390)>sat (922)=cis mono (869)=n-6 poly (868)>n-3 poly (652)>commercial diet (526). Significant elevations in very low density lipoprotein cholesterol were also found in the trans and cis mono and sat groups, and triacylglycerol concentrations were also elevated in all groups. High density lipoprotein cholesterol concentrations were consistently low (20 to 50 mg/dL) in all groups. Highly significant differences (ANOVA, P<0.0001) in atherosclerosis, quantified by measurement of aortic cholesteryl ester concentration (mg/g protein) among dietary fatty acid groups were found, with the order being trans mono (mean, 50.4)>sat (35.6)=cis mono (34.6)>n-6 poly (18. 3)=n-3 poly (9.7)=commercial diet (7.8). Therefore, in this mouse model of hypercholesterolemia, dietary cis or trans monounsaturated fat did not protect against atherosclerosis development, whereas aortic atherosclerosis in either of the polyunsaturated fat groups was significantly less than in the saturated fat group.     

Headaches in persons attending polyclinics in Barbados

Assessment and documentation of patient responses to treatment and/or invasive procedures are important facets of nursing care. The purpose of this article is to describe a tool for assessing and quantifying a patient's response to sedation and tolerance of a specific diagnostic and/or therapeutic procedure. Use of the conscious sedation scale will allow the nurse to objectively measure the patient's outcome. This tool will also facilitate better patient care and documentation and substantiate nursing intervention.     

Absence of antinociceptive tolerance to improgan, a cimetidine analog, in rats

Improgan, an analog of the histamine receptor antagonist cimetidine, produces highly effective analgesia following intraventricular injection. The present study examined changes in the antinociceptive effects of improgan following once daily intraventricular injections. Improgan (100-150 microg) produced near maximal antinociception 10 and 30 min after daily administration on all 4 test days, whereas comparable morphine treatments (50 microg) induced considerable tolerance. Thus, improgan produced highly effective analgesia without the development of tolerance.