Display of the alveolar plateau of single-breath tests in "dilution index" format

In 1949, Fowler (J. Appl. Physiol. 2: 283-299) advocated calculation of a "dilution index" from data of the alveolar plateau of single-breath tests; the calculation provides an estimate of the dilution of resident gas in the lung that gave rise to the observed concentrations. In this communication, we show that the calculation can be applied to conventional single-breath tests where O2 is inhaled by air-breathing persons, and we illustrate the principle with vital capacity breaths of a mixture that contained a low concentration of neon. The dilution was approximately 3:1 in young subjects (20-30 yr), as if a vital capacity of 6 liters were mixed with a residual volume of 2 liters. The dilution was less, 2:1, in older subjects (56 yr) and tended to become as low as 1:1 during emptying of the closing volume. In addition to being more informative, the dilution index format allows common sense comparison of alveolar plateau levels and slopes when single-breath tests are done by various methods.     

Suppressive substance produced by T cells from mice chronically infected with Trypanosoma cruzi. I. Preferential inhibition of the induction of delayed-type hypersensitivity

Culture supernatants of spleen cells from susceptible CBA mice chronically infected with Trypanosoma cruzi were able to inhibit the induction of delayed-type hypersensitivity (DTH) to a wide range of antigens as measured by 24-hr footpad swelling, bone marrow homing, and radioactivity accumulation assays. The suppressive activity, which was also present in the serum of these chronically infected mice, appears to be specific for the induction of DTH and had no effect on the 3-hr immediate-type hypersensitivity. It also failed to modify the expression of DTH in presensitized mice. Furthermore, it did not affect the synthesis in normal recipients of specific antibody or the induction of helper T cells or cytotoxic T cells. It also failed to induce DTH tolerance as recipient mice with markedly reduced DTH were able to develop a normal DTH response after secondary immunization. The suppressive activity was produced by an Ig- macrophage-depleted splenic T cell population, whose capacity to secrete the suppressive substance was completely abrogated by treatment in vitro with anti-L3T4 antibody and complement, but not with anti-Lyt-2 antibody and complement. These results therefore demonstrate that L3T4+ T cells from mice chronically infected with T. cruzi can produce substances which interfere with the induction of DTH. This finding may help to identify the differential antigenic stimulatory requirement for the activation of the various subsets of T cells.     

The estimation of host antigen in experimental and commercial influenza subunit preparations by delayed-type hypersensitivity reaction

A delayed-type hypersensitivity (DTH) test has been used to detect the presence of host-specific antigen(s) in highly-purified egg-grown influenza virus and virus components. Cyclophosphamide-treated mice, primed with an extract from allantoic fluid, showed DTH reactions when challenged with whole virus, spikeless virus particles or polymer-type subunits. All commercial egg-grown influenza vaccines showed the presence of host antigen, whereas lipid-free derivatives such as internal virus protein complexes and monomer HA preparations lacked the activity. These findings are discussed in relation to vaccine application.     

Interleukin-10 (IL-10) inhibits the induction of nitric oxide synthase by interferon-gamma in murine macrophages.

A murine macrophage cell line, J774, expresses high levels of the enzyme nitric oxide synthase (NOS) and produces large amounts of nitric oxide (NO) when activated with recombinant interferon (IFN)-gamma and a low concentration of LPS (10 ng/ml). Both the expression of NOS and the production of NO were inhibited by recombinant IL-10 in a dose-dependent manner. The inhibition was effective only when the cells were pretreated with IL-10; addition of IL-10 at the same time or after IFN-gamma activation was without effect. These results demonstrate that IL-10, a product of Th2 (helper T lymphocyte 2) cells, can antagonise the function of IFN-gamma, a product of Th1 cells, by modulating the mechanism of synthesis of nitric oxide in the macrophages.     

Regulation of transglutaminase 1 gene expression by 12-O-tetradecanoylphorbol-13-acetate, dexamethasone, and retinoic acid in cultured human keratinocytes.

Transglutaminase 1 (TG1) is an enzyme that is expressed at the late stage of terminal differentiation of keratinocytes and catalyzes the epsilon-(gamma-glutamyl)lysine cross-linking reaction to form a highly insoluble cell envelope. To elucidate the mechanism of TG1 gene expression in keratinocytes, we examined the effects of 12-O-tetradecanoylphorbol-13-acetate (TPA), dexamethasone, 1,25-dihydroxyvitamin D3, and retinoic acid on the levels of TG1 mRNA in cultured normal human epidermal keratinocytes (NHEK). Treatment of NHEK with TPA, up to 10 nM, markedly increased the levels of TG1 mRNA in a dose-dependent manner. The effect by treatment with 1 nM TPA reached a peak after 16 h of incubation (20-fold above the basal level). In contrast, phorbol had no effect on TG1 gene expression. The induction of TG1 mRNA expression by TPA was inhibited by 1-(5-isoquinolinylsulfonyl)-2-methyl-piperazine (H-7) and staurosporine. Dexamethasone at a concentration of 1 microM also increased the TG1 mRNA levels, but the maximum induction was observed (3-fold above the basal level) after 72 h of incubation. The effect of dexamethasone was not suppressed by H-7. Moreover, 1 microM of retinoic acid completely inhibited the induction of TG1 mRNA by both TPA and dexamethasone. 1,25-Dihydroxyvitamin D3 showed no effect on the TG1 mRNA levels. From these results, we suggest that the expression of TG1 gene may be upregulated by protein kinase C and glucocorticoid receptor systems and down-regulated by the retinoic acid receptor system.     

Two different forms of beta myosin heavy chain are expressed in human striated muscle

Summary We have found evidence for two beta-like myosin heavy chains in humans, one cardiac and one skeletal. The cDNA sequences of the cardiac beta myosin heavy chain cDNA clone pHMC3 and the skeletal beta-like myosin heavy chain cDNA clone pSMHCZ, were compared to each other. It was found that the 3 untranslated regions as well as 482 nucleotides specifying the carboxyl coding region, were 100% homologous. Further examination revealed that the skeletal clone pSMHCZ diverges from the human cardiac beta myosin heavy chain cDNA clone pHMC3 at the 5 end. We present evidence in this report which indicates that the cardiac beta myosin heavy chain mRNA is expressed in skeletal muscle tissues. The human cardiac beta myosin heavy chain cDNA clone, pHMC3, which codes for a portion of the light meromyosin section of the myosin heavy chain, was used as a probe for S1 nuclease mapping studies with RNA derived from cardiac tissue, smooth muscle and skeletal muscle tissues consisting of fast-twitch, slow-twitch and mixed fast- and slow-twitch muscle fibres. Two probes were used to examine the expression of the mRNA. One probe (406 nucleotides) constitutes the 3 untranslated region and a portion of the coding region of the beta cardiac myosin heavy chain cDNA clone, which is 100% homologous to pSMHCZ, the skeletal cDNA clone. The other constitutes the majority of the coding region (1017 nucleotides) of the cardiac clone pHMC3 in which the first 216 nucleotides from the labelled end are 100% homologous to the skeletal clone pSMHCZ. In the soleus muscle, which is rich in slow-twitch type I muscle fibres, the expression of the cardiac beta myosin heavy chain mRNA was very prominent. In gastrocnemius muscle, a mixed fibre muscle, the expression of this mRNA was detected to a lesser degree than that for the soleus muscle. In vastus lateralis and vastus medialis, which consist of predominantly type II, fast-twitch fibres, there were trace amounts of the cardiac beta myosin heavy chain mRNA. When expression of this mRNA was tested in smooth muscle tissue none could be detected.     

Thermostabilization of enteroviruses by estuarine sediment

The effect of estuarine sediment on the thermoinactivation of poliovirus type 1 and echovirus type 1 was evaluated. Poliovirus survival was prolonged at 24 and 37 degrees C but not at 4 degrees C in the presence of sediment over the time periods observed. Further inactivation studies were performed at 50 and 55 degrees C to maximize the thermal effects, and similar protection was observed. The supernatant fluid from a mixture of seawater and sediment lacked the protective effect against thermoinactivation, suggesting that prolonged virus survival in the presence of sediment was due to adsorption to particulates. From these observations, it appears that the adsorption of enteroviruses to estuarine sediments may play a significant role in protecting them against thermoinactivation.     

Construction of cosmid genomic libraries for the normal and myopathic Syrian hamsters

Cosmid genomic libraries from both normal and myopathic Syrian hamsters have been constructed. MboI was used to generate 35- to 50-kilobase DNA fragments which were isolated from a 5-25% NaCl gradient. The 35- to 50-kilobase DNA fragments were ligated to the cosmid vector pCV108 and packaged into Escherichia coli DK1. Approximately 3 X 10(5) - 4 X 10(5) clones were obtained per microgram of ligated DNA. Thirteen clones have been isolated from 2 X 10(5) colonies using a cardiac myosin heavy chain clone as a probe. Restriction maps of two of these clones are presented here.