Biomedical subjects

L M Lee

Publications and source records attributed to L M Lee.

At least 19 recordsLinked to original sources

PubMed2006

Lipoprotein lipase gene S447X polymorphism modulates the relation between central obesity and serum lipids, a twin study.

OBJECTIVE: To investigate the impact of lipoprotein lipase (LPL) gene S447X polymorphism on the relation between central obesity and lipid levels. DESIGN: A total of 961 adult twin pairs were included from the program of Chinese Twin Registry, between 2001 and 2002. Central obesity was defined as waist circumference > or =90 cm for male and > or =80 cm for female. Two statistical methods were performed to test the modification effect of S447X polymorphism of LPL gene on the relation between central obesity and lipid levels: one was Generalized Estimating Equation (GEE) models for all twin pairs and the other was co-twin matched case-control analysis in 82 central obesity discordant monozygotic twin pairs. RESULTS: In GEE models for all twins, central obesity was significantly associated with serum lipids except for high-density lipoprotein (HDL), while X447 allele had favorable effects on the levels of triglyceride (TG), HDL and TG to HDL ratio (TG/HDL). The interactions of S447X polymorphism and central obesity were statistically significant for TG/HDL and HDL. In central obesity discordant monozygotic twin pairs, central obesity was significantly related with 26.2% increase of TG and 27.2% increase of TG/HDL in S/S447 genotype, while in 447X allele carrier, central obesity was significantly related with 13.7% increase of HDL. CONCLUSION: These results suggest that LPL gene S447X polymorphism modifies the relation between central obesity and serum lipids, which also stresses the importance of reducing waist circumference to improve serum lipids for people with central obesity, especially those with S/S447 genotype.

Adult↗

PubMed2006

Low adherence to guidelines for preventing TB among persons with newly diagnosed HIV infection, United States.

SETTING: Persons infected with human immunodeficiency virus (HIV) are at risk for developing tuberculosis (TB) if latent TB infection remains untreated. OBJECTIVE: To assess missed opportunities for preventing TB by selecting a population-based sample of 1093 persons diagnosed with HIV from June 1995 to June 1997 in Seattle, WA, New Orleans, LA, and Jersey City, NJ. DESIGN: To determine the proportion of persons receiving a tuberculin skin test (TST) following HIV diagnosis, we conducted record reviews at providers and local TB control. RESULTS: An estimated 53.7% (95% CI 49.9-57.4) had a TST following HIV diagnosis; 6.6% (95% CI 4.3-8.9%) of TST-tested patients were reactive. Median time between HIV diagnosis and TST was 1 month (mean 5.7 months, 95% CI 4.8-6.5). Factors associated with TST included additional risk factors for TB (OR 1.76, 95% CI 1.17-2.63), history of HIV-related preventive treatment (OR 5.84, 95% CI 3.74-8.75), higher number of clinic visits (OR 4.16, 95% CI 2.01-8.02), and attendance at facilities with a written policy to provide TST for all persons with HIV (OR 2.54, 95% CI 1.28-4.88). CONCLUSION: About half of persons newly diagnosed with HIV infection had a TST following HIV diagnosis, with little variation by demographics, signaling a general need to improve interventions to prevent TB.

Female↗

PubMed2005

Latent membrane protein 1 encoded by Epstein-Barr virus modulates directly and synchronously cyclin D1 and p16 by newly forming a c-Jun/Jun B heterodimer in nasopharyngeal carcinoma cell line.

Recently we confirmed that latent membrane protein 1 (LMP1) encoded by Epstein-Barr virus (EBV) accelerates a newly forming active c-Jun/Jun B heterodimer, a transcription factor, but little is known about the target gene regulated by it. In this paper, results indicated that a c-Jun/Jun B heterodimer induced by LMP1 upregulated cyclin D1 promoters activity and expression, on the contrary, downregulated p16, and maladjustment of cyclin D1 and p16 expression accelerated progression of cell cycle. Firstly, we found a c-Jun/Jun B heterodimer regulated synchronously and directly cyclin D1 and p16 in the Tet-on-LMP1-HNE2 cell line, in which LMP1 expression is regulated by Tet-on system. This paper investigated in depth function of the newly forming active c-Jun/Jun B heterodimer, and built new connection between environmental pathogenic factor, signal transduction and cell cycle.

Carcinoma↗

PubMed2005

Cotton bud and ear cleaning--a loose tip cotton bud?

Complications that arise from self-cleaning the external ear canal are common. This is a prospective study using standard questionnaire on the practice of ear cleaning in 50 subjects. A loose tip cotton bud was also shown to them to assess if it was acceptable to replace their current cotton bud. Thirty six percent of them clean their ears by introducing an object into the ear canal once or more a day. The commonest reason to clean the ears was the presence of earwax. Only 6% used a cotton bud to mop fluid from the ear canal. Almost all (92%) the subjects used a cotton bud to clean the ear. A complication rate of 2% was noted. A loose tip cotton swab was shown to the subjects. Despite explaining that it is safer and better, only 24% of the subject were willing to change to this loose tip cotton bud. Seventy four percent of the subjects cleaned their ear regularly because of earwax. The misconception of needing to clean the ear canal by introducing an object into the ear is rampant. This practice should be avoided and health care providers can play an important role in this.

Adolescent↗

PubMed2003

Pre and post treatment mucociliary function in allergic rhinitis in three different treatment modalities.

Allergic rhinitis causes an impairment of the mucociliary function in the nose. It is hoped that treatment of perennial allergic rhinitis would be able to revert mucociliary function to normal. This study aims to compare pre and post treatment mucociliary transport time in 3 different treatment modalities. Ninety-two newly diagnosed patients with allergic rhinitis were randomised into 3 groups and started on different treatment regimes. At the end of 8 weeks, the group treated with only intranasal beclomethasone showed some, though not significant, improvement in the mucociliary function. There were no changes in the mucociliary function in the other two groups treated with beclomethasone and loratidine or loratidine alone.

Administration, Intranasal↗

PubMed2002

Drug-drug interactions in inmates treated for human immunodeficiency virus and Mycobacterium tuberculosis infection or disease: an institutional tuberculosis outbreak.

The use of rifamycins is limited by drug interactions in human immunodeficiency virus (HIV)-infected persons who are receiving highly active antiretroviral therapy (HAART). During a tuberculosis (TB) outbreak at a prison housing HIV-infected inmates, rifabutin was used to treat 238 men (13 case patients and 225 contacts). Steady-state peak plasma rifabutin concentrations were obtained after rifabutin dosages were adjusted for men receiving single-interacting HAART (with either 1 protease inhibitor [PI] or efavirenz), multi-interacting HAART (with either 2 PIs or > or =1 PI with efavirenz), and for noninteracting HAART (>1 nucleoside reverse-transcriptase inhibitor or no HAART) without rifabutin dose adjustments. Low rifabutin concentrations occurred in 9% of those receiving noninteracting HAART, compared with 19% of those receiving single-interacting and 29% of those receiving multi-interacting HAART (chi2, 3.76; P=.05). Of 225 contacts treated with rifabutin-pyrazinamide, 158 (70%) completed treatment while incarcerated. Rifabutin-pyrazinamide therapy was difficult to implement, because of the need for dosage adjustments and expert clinical management.

Adult↗

PubMed2001

Survival after AIDS diagnosis in adolescents and adults during the treatment era, United States, 1984-1997.

CONTEXT: Declines in the number of acquired immunodeficiency syndrome (AIDS) deaths were first observed in 1996, attributed to improvements in antiretroviral therapy and an increase in the proportion of persons receiving therapy. OBJECTIVE: To examine national trends in survival time among persons diagnosed as having AIDS in 1984-1997. DESIGN, SETTING, AND SUBJECTS: Retrospective cohort study using data from a population-based registry of AIDS cases and deaths reported in the United States. MAIN OUTCOME MEASURE: Months of survival after AIDS diagnosis through December 31, 1998, compared by year of diagnosis. RESULTS: Among 394 705 persons with an AIDS-defining opportunistic illness (OI) diagnosed in 1984-1997, median survival time improved from 11 months for 1984 diagnoses to 46 months for 1995 diagnoses. Among persons with an OI diagnosed in 1996 and 1997, 67% were alive at least 36 months after diagnosis and 77% were alive at least 24 months after diagnosis, respectively. Among 296 621 AIDS cases diagnosed during 1993-1997, 65% were based on immunologic criteria and 35% on OI criteria; 80% were among men; and 42% were among non-Hispanic blacks, 40% among non-Hispanic whites, 17% among Hispanics, 1% among Asians/Pacific islanders, and less than 1% among American Indians/Alaska natives. The probability of surviving at least 24 months increased from 67% for those with immunologic diagnoses in 1993 to 90% in 1997 and from 49% for those with OI diagnoses in 1993 to 80% in 1997. Survival time increased with each year of diagnosis from 1984 to 1997 for blacks, whites, and Hispanics. The greatest annual survival gains occurred among persons receiving an AIDS diagnosis in 1995 and 1996. CONCLUSIONS: Survival time after AIDS diagnosis improved from 1984 to 1997. While AIDS incidence is declining, improved survival times present a growing public health challenge as the number of persons living with chronic human immunodeficiency virus disease/AIDS increases.

AIDS-Related Opportunistic Infections↗

PubMed2001

Emerging viral infections.

The past decade has witnessed the emergence of several significant viral pathogens and the further evolution of additional viral pathogens. Transmitted by a variety of differing routes, these organisms have presented substantial intellectual challenges to medicine of the 20th and 21st centuries. As perhaps the benchmark pathogen of the past decade, HIV has provided medicine and society with a most formidable opponent, and one that has yet to be fully conquered. Nonetheless, a variety of additional viral pathogens have also perplexed medicine over the past 10-15 years.

Communicable Diseases, Emerging↗

PubMed2001

Low molecular weight protamine (LMWP) as nontoxic heparin/low molecular weight heparin antidote (II): in vitro evaluation of efficacy and toxicity.

Patients undergoing anticoagulation with heparin or low molecular weight heparin (LMWH) require a superior antidote that possesses more selective biological actions and a better safety profile than protamine. We had previously developed 2 low molecular weight protamine (LMWP) fractions (TDSP4 and TDSP5) from thermolysin-digested protamine as potential nontoxic, heparin-neutralizing agents. In this, the second article in this series, studies focused on in vitro evaluation of heparin/LMWH-neutralizing efficacy and putative toxicity. These LMWP fractions, particularly TDSP5, were effective and fully capable of neutralizing a broad spectrum of heparin-induced anticoagulant activities (ie, aPTT, anti-Xa, and anti-IIa activities). Additionally, these LMWP fractions could neutralize the activities of commercial LMWH. As assessed by the anti-Xa assay, TDSP5 was as effective as, although less potent than, protamine in reversing the activity of Mono-Embolex (molecular weight 5000-7000) and 2 other different sizes (molecular weight of 3000 and 5000 d) of LMWH preparations. Furthermore, compared with protamine, TDSP5 exhibited a much-reduced toxicity and thus an improved safety profile, as reflected by its reduced ability to activate the complement system and cross-react with the antiprotamine antibodies, which are 2 primary indices of protamine toxicity.

Animals↗

PubMed2001

Low molecular weight protamine as nontoxic heparin/low molecular weight heparin antidote (III): preliminary in vivo evaluation of efficacy and toxicity using a canine model.

Heparin employed in cardiovascular surgeries often leads to a high incidence of bleeding complications. Protamine employed in heparin reversal, however, can cause severe adverse reactions. In an attempt to address this clinical problem, we developed low molecular weight protamine (LMWP) as a potentially effective and less toxic heparin antagonist. A homogeneous 1880-d peptide fragment, termed LMWP-TDSP5 and containing the amino acid sequence of VSRRRRRRGGRRRR, was derived directly from protamine by enzymatic digestion of protamine with thermolysin. In vitro studies demonstrated that TDSP5 was capable of neutralizing various anticoagulant functions of both heparin and commercial low molecular weight heparin preparations. In addition, TDSP5 exhibited significantly reduced crossreactivity toward mouse sera containing antiprotamine antibodies. TDSP5 showed a decrease in its potential in activating the complement system. All of these findings suggested the possibility of markedly reduced protamine toxicity for TDSP5. In this article, we conducted preliminary in vivo studies to further demonstrate the feasibility and utility of using LMWP as a nontoxic clinical protamine substitute. Dogs were chosen as test animals because they were known to magnify the typical human response to protamine. By using a full spectra of biological and clinical assays for heparin, including the anti-IIa and anti-Xa chromogenic assays and the activated partial, thromboplastin time and TCT clotting assays, TDSP5 showed that it could completely neutralize all these different anticoagulant functions of heparin in dogs. Although administration of protamine in dogs produced a significant reduction in mean arterial blood pressure (-14.9 mm Hg) and elevation in pulmonary artery systolic pressure (+5.0 mm Hg), the use of TDSP5 in dogs did not elicit any statistically significant change in any of the variables measured. Furthermore, the use of LMWP also significantly reduced the protamine-induced transient thrombocytopenic and granulocytopenic responses. The white blood cell counts and platelet counts decreased to 82.1% and 60.0% of baseline, respectively, in dogs given intravenous protamine compared to 97.8% and 88.6% of baseline in dogs receiving TDSP5. These preliminary findings indicated that LMWP could potentially provide an effective and safe means to control both heparin- and protamine-induced complications.

Animals↗

PubMed2001

Effector genes altered in MCF-7 human breast cancer cells after exposure to fractionated ionizing radiation.

Understanding the molecular mechanisms involved in the response of tumors to fractionated exposures to ionizing radiation is important for improving radiotherapy and/or radiochemotherapy. In the present study, we examined the expression of stress-related genes in an MCF-7 cell population (MCF-IR20) that has been derived through treatment with fractionated irradiation (2 Gy per fraction with a total dose of 40 Gy). MCF-IR20 cells showed a 1.6-fold increase in sensitization with dose at 10% isosurvival in a clonogenic assay, and a reduced growth delay ( approximately 15 h compared to approximately 27 h), compared to the parental MCF-7 cells treated with a single dose of 5 Gy. To determine which effector genes were altered in the MCF-IR20 cells, the expression of stress-related effector genes was measured using a filter with 588 genes (Clontech) that included major elements involved in cell cycle control, DNA repair, and apoptosis. Compared to MCF-7 cells that were not exposed to fractionated radiation, 19 genes were up- regulated (2.2-5.1-fold) and 4 were down-regulated (2.7-3.4- fold) in the MCF-IR20 cells. In agreement with the array results, 6 up-regulated genes tested by RT-PCR showed elevated expression. Also, activities of the stress-related transcription factors NFKB, TP53 and AP1 showed a 1.2-4.5-fold increase after a single dose of 5 Gy in MCF-IR20 cells compared with parental MCF-7 cells. However, when the radioresistant MCF-IR20 cell were cultured for more than 12 passages after fractionated irradiation (MCF-RV), radioresistance was lost, with the radiosensitivity being the same as the parental MCF- 7 cells. Interestingly, expression levels of CCNB1, CD9 and CDKN1A in MCF-RV cells returned to levels expressed by the parental cells, whereas the expression levels of three other genes, MSH2, MSH6 and RPA remained elevated. To determine if any of the changes in gene expression could be responsible for the induced radioresistance, CCNB1 and CDKN1A, both of which were up-regulated in MCF-IR20 cells and down-regulated in MCF-RV cells, were studied further by transfection with antisense oligonucleotides. Antisense of CCNB1 significantly reduced the clonogenic survival of MCF- IR20 cells at doses of 5 and 10 Gy, from 42% to 26% and from 5.7% to 1.0%, respectively. Antisense of CDKN1A, however, had no effect on radiation survival of MCF-IR20 cells. In summary, these results suggest that stress-related effector genes are altered in cells after treatment with fractionated irradiation, and that up-regulation of CCNB1 is responsible, at least in part, for radioresistance after fractionated irradiation.

Adenocarcinoma↗

PubMed2001

Tolerability of postexposure antiretroviral prophylaxis for occupational exposures to HIV.

A substantial body of evidence provides support (but not definitive proof of efficacy) for the use of antiretroviral agents as postexposure prophylaxis for occupational exposures to HIV in the healthcare workplace. Despite the lack of definitive evidence of the efficacy of these agents in this setting, over the past decade this intervention has become the standard of care for healthcare workers who sustain occupational exposures to HIV. Administration of these agents--even for a relatively short 28-day postexposure course--is often fraught with difficulty. All of the agents currently used for postexposure prophylaxis regimens have substantial adverse effects, and significant adverse effects occur in more than two-thirds of individuals electing prophylaxis. This manuscript reiterates current US Federal Government guidelines for the administration of postexposure prophylaxis, specifically noting that zidovudine plus lamivudine (with or without a protease inhibitor) remains the recommended regimen. The paper summarises the significant toxicities associated with nucleoside reverse transcriptase inhibitors (primarily nausea, vomiting, diarrhoea and bone marrow suppression), non-nucleoside reverse transcriptase inhibitors (rash, fever, gastrointestinal symptoms and hepatitis, including hepatic decompensation necessitating liver transplantation) and protease inhibitors (nausea, vomiting, diarrhoea, abdominal pain, hyperglycaemia, hyperlipidaemia, headache and anorexia). As a class, the antiretroviral agents have an extraordinary number of drug interactions. The non-nucleoside reverse transcriptase inhibitors and the protease inhibitors are metabolised through the cytochrome P450 pathway, and the effects of concomitant administration of protease inhibitors with other agents in the same class are discussed, as well as the effects of concomitant administration of protease inhibitors with non-nucleoside agents. The potential for numerous and medically risky drug interactions emphasises the importance of planning antiretroviral prophylaxis in consultation with practitioners or clinical pharmacists who are skilled in the use of these agents and knowledgeable about the potential for significant drug interactions that could either reduce the benefit of prophylaxis or increase the potential for toxicity. Another common problem encountered by individuals managing postexposure prophylaxis programmes relates to the administration of chemoprophylaxis to a pregnant healthcare worker who has sustained an occupational exposure to HIV. We address what is known about the potential for toxicity and emphasise the recently published warning concerning the deaths of pregnant women and their offspring from lactic acidosis while receiving regimens containing stavudine and didanosine.

Anti-HIV Agents↗

PubMed2001

Trends in human immunodeficiency virus diagnoses among women in the United States, 1994-1998.

OBJECTIVE: Since the introduction of combination antiretroviral therapy in 1996, acquired immune deficiency syndrome (AIDS) data no longer provide information about trends in human immunodeficiency virus (HIV) incidence; therefore, we used HIV surveillance data to describe recent trends in HIV diagnoses among women. METHODS: Using HIV cases reported from 25 states with confidential HIV reporting, we examined new HIV diagnoses from 1994 to 1998 among women born between 1950 and 1979 by five-year birth cohorts. We adjusted for reporting delays and anticipated reclassification of cases reported without exposure risk. RESULTS: During the period, 24 171 cases of HIV infection were diagnosed among women. The annual number increased 4% from 1994 to 1995, then declined 12% from 1995 to 1998. The annual number of diagnoses attributed to heterosexual contact (HC) among women age 15 to 19 in 1994 (born 1975-1979) increased 117% from 1994 to 1998. Among older women, HC-related diagnoses remained stable or declined. The annual number of diagnoses attributed to injection drug use (IDU) among women age 15 to 19 in 1994 increased 90% from 1994 to 1998. The number of IDU-related diagnoses among older women declined between 31% and 59%. Diagnosis rates were higher among HC-related cases than among IDU cases. The rate for IDU-related diagnoses declined by nearly half over the period, but remained stable for HC-related diagnoses. CONCLUSIONS: HIV diagnoses among women decreased slightly between 1994 and 1998. As the youngest cohort of women reached the age where risk behaviors are initiated, however, the number of diagnoses attributed to IDU increased and the number of diagnoses attributed to HC more than doubled. It is in this youngest cohort, where risk behaviors have only recently been initiated, that HIV diagnoses most closely approximate trends in HIV incidence.

Adolescent↗

PubMed2000

Duration of human immunodeficiency virus infection and likelihood of giving birth in a Medicaid population in Maryland.

The objective of this study was to examine the effect of duration of human immunodeficiency virus (HIV) infection on a woman's likelihood of giving birth. Using longitudinal data from the Maryland state Human Immunodeficiency Virus Information System and a retrospective cohort design, the authors compared 1,642 women with acquired immunodeficiency syndrome (AIDS) to 8,443 uninfected women enrolled in the Medicaid program between 1985 and 1995. The decade before AIDS diagnosis was divided into four 2.5-year periods. Proximity to AIDS diagnosis served as a proxy for duration of infection. An extension of the Cox model was used to estimate the relative risk for giving birth, with adjustment for covariates and repeated outcomes. The average number of births per 100 person-years was 6.0 for HIV-infected women and 11.1 for uninfected women (adjusted relative risk = 0.63; 95% confidence interval (CI): 0.57, 0.68). Accounting for duration of infection, the adjusted relative risks for birth among HIV-infected women, as compared with uninfected women, were 0.85 (95% CI: 0.71, 1.03), 0.74 (95% CI: 0.63, 0.86), 0.55 (95% CI: 0.47, 0.64), and 0.45 (95% CI: 0.38, 0.55) for successive 2.5-year periods before AIDS diagnosis. Demographic characteristics, contraception, abortion, fetal loss, or drug use could not fully explain the reductions. These results suggest that HIV-infected women experience a progressive reduction in births years before the onset of AIDS. This may compromise estimation of HIV prevalence and interpretation of time trends from serosurveillance of pregnant women.

Adolescent↗

PubMed2000

Mutagenesis reveals a role for ABP/GRIP binding to GluR2 in synaptic surface accumulation of the AMPA receptor.

We studied the role of PDZ proteins GRIP, ABP, and PICK1 in GluR2 AMPA receptor trafficking. An epitope-tagged MycGluR2 subunit, when expressed in hippocampal cultured neurons, was specifically targeted to the synaptic surface. With the mutant MycGluR2delta1-10, which lacks the PDZ binding site, the overall dendritic intracellular transport and the synaptic surface targeting were not affected. However, over time, Myc-GluR2delta1-10 accumulated at synapses significantly less than MycGluR2. Notably, a single residue substitution, S880A, which blocks binding to ABP/GRIP but not to PICK1, reduced synaptic accumulation to the same extent as the PDZ site truncation. We conclude that the association of GluR2 with ABP and/or GRIP but not PICK1 is essential for maintaining the synaptic surface accumulation of the receptor, possibly by limiting its endocytotic rate.

Adaptor Proteins, Signal Transducing↗

PubMed2000

Effects of handicap on life expectancy: the case of China.

The purpose of this study was to quantify and partition the expected years of life with and without handicap for the Chinese population according to various types of handicaps, age-sex groups and regions. A large-scale sample survey on handicapped persons conducted in 1987, and the 1990 population census constitute the basis for computing the expected years of life free of handicapped condition using the method proposed by Sullivan. The expected years of life with handicap for the Chinese population in childhood (0-14 y), working ages (15-64 y) and the elderly (65 y+) were 0.40, 1.78, and 3.44 for males and 0.34, 1.69, and 4.55 for females. For the Chinese males over 65 y of age, there were about 1.83 expected years of life with aural handicap and 0.59 expected years of life with ocular handicap. For the Chinese females over 65 y of age, there were about 1.87 expected years of life with aural handicap and 1.16 expected years of life with ocular handicap. The burden of living with handicap is greater for females and the elderly. This general pattern hold for all types of handicap except for skeletal handicap. The expected years of life with handicap for the Chinese population provide useful information for setting public health policies, despite the difficulty in making comparisons with the similar data in other countries.

Adolescent↗

PubMed1998

The AMPA receptor GluR2 C terminus can mediate a reversible, ATP-dependent interaction with NSF and alpha- and beta-SNAPs.

In this study, we demonstrate specific interaction of the GluR2 alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptor subunit C-terminal peptide with an ATPase N-ethylmaleimide-sensitive fusion protein (NSF) and alpha- and beta-soluble NSF attachment proteins (SNAPs), as well as dendritic colocalization of these proteins. The assembly of the GluR2-NSF-SNAP complex is ATP hydrolysis reversible and resembles the binding of NSF and SNAP with the SNAP receptor (SNARE) membrane fusion apparatus. We provide evidence that the molar ratio of NSF to SNAP in the GluR2-NSF-SNAP complex is similar to that of the t-SNARE syntaxin-NSF-SNAP complex. NSF is known to disassemble the SNARE protein complex in a chaperone-like interaction driven by ATP hydrolysis. We propose a model in which NSF functions as a chaperone in the molecular processing of the AMPA receptor.

Adenosine Triphosphate↗

PubMed1997

A note on the preparation of whole mount samples suitable for observation with the confocal laser scanning microscope.

The effects of fixatives and pretreatment on the immunofluorescence of whole mount specimens prepared for confocal laser scanning microscopy (CLSM) were examined. Intact villi were obtained from the proximal small intestine of mice fixed with 4% paraformaldehyde (4P) or 0.5% paraformaldehyde and 15% of saturated picric acid (PPa). Before immunostaining for laminin and tenascin, each specimen was pretreated with deoxycholate, while some 4P-fixed specimens received further pepsin pretreatment. Regardless of the fixatives employed, laminin and tenascin showed adequate immunofluorescence. Without pepsin pretreatment, the 4P-fixed specimens emitted conspicuous background fluorescence, and immunofluorescence was weak in the lamina propria. Pepsin pretreatment reduced the background fluorescence, but also diminished the immunofluorescence, especially that of tenascin. The PPa-fixed specimens displayed intense immunofluorescence of laminin and tenascin with very little background, even deeply within the lamina propria. When the PPa-fixed specimens were immunostained for vasoactive intestinal peptide, immunopositive nerve fibres were observed within the lamina propria. Ultrastructural investigation of the PPa-fixed specimens revealed that membranous structures in all cells were almost lost while tissue architecture was well preserved. These results indicate that PPa fixation and pretreatment with deoxycholate are suitable for preparing whole mount specimens for CLSM studies.

Animals↗