Lymphocyte subpopulations in healthy 1-3-day-old infants

The primary objective of this study was to establish reference ranges for the major (B, T, and natural killer; NK) and clinically relevant minor lymphocyte subsets in the peripheral blood of healthy 1-3-day-old infants and then to compare the results with those obtained in a group of healthy adults analyzed simultaneously. Forty-three infants aged 1-3 days and 38 healthy adults were recruited to the study to establish the median, 10th, and 90th percentiles of the proportions and absolute numbers of relevant lymphocyte subsets. The samples obtained from the healthy adults served as a flow cytometry process control in addition to providing a group comparator. The peripheral blood of the newborns (vs. adults) contained elevated proportions of total T cells (83% vs. 77%) and T helper cells (63% vs. 46%), with decreased proportions of T suppressor/cytotoxic cells (23% vs. 28%) and NK cells (4% vs. 10.5%). The newborns had a higher proportion (P < 0.0001) of immature B lymphocytes compared with those of adults (CD10+CD19+, 1.5% vs. 0% and CD20+CD5+, 13% vs. 6%), and the proportion of activated T cells was significantly lower (P < 0.0001; CD3+CD25+, 7.0% vs. 15%;CD3+HLA-DR+, 2.0% vs. 6% and CD8 and CD57, 0.0% vs. 8.0%). In contrast, the proportions of neonatal CD8 cells expressing CD28 (90.2% vs. 67.7%) and CD38 (96.6% vs. 70.9%) were significantly higher (P < 0.0001). The reference ranges for 1-3-day-old healthy newborns generated in this study provides a valuable tool for the assessment of immune abnormalities in very young infants.     

Estrogen receptors and the mammary gland

For several decades it has been known that steroid hormones, estrogen and progesterone, regulate some genes involved in the growth, proliferation and differentiation of the mammary-gland in animals and humans. In the last years, the presence or absence of the nuclear estrogen receptor has been used by clinicians as a marker for tumor malignancy, as a prognostic index or as an important parameter for hormonal therapy with anti-estrogenic compounds of some hormone-dependent breast cancers. This review shows some advances in the knowledge of the structure, function, molecular mechanisms of estrogenic activity, and interaction with proteins like protooncogenes and growth factors. Also, we refer to the role of the estrogen receptor in the physiophatology of breast cancer.     

Identification of lactoferrin complexes in bovine mammary secretions during mammary gland involution

Part of the antimicrobial activity of lactoferrin resides in its ability to bind to bacteria. The complexing of lactoferrin with other proteins could alter its activity. This study identified the presence of lactoferrin complexes in mammary secretions during mammary gland involution and determined the proportion of free and complexed lactoferrin in mammary secretions. Mammary secretions were collected from Holstein cows on d 7, 14, and 21 of involution. Proteins were fractionated from defatted, filtered mammary secretions by sucrose density gradient ultracentrifugation and by gel filtration chromatography. Proteins contained in separated fractions were identified by SDS-PAGE. The presence of lactoferrin was confirmed by immunoblot analysis. Lactoferrin was present as complexed forms of high molecular mass in mammary secretions at each day of involution. The majority of lactoferrin was present in complexes of higher molecular mass rather than as monomers. A majority of lactoferrin existed in fractions of approximately 250 kDa, although peaks of lactoferrin at 150, 300, and 800 kDa were also found. The presence of lactoferrin complexes may result from interactions with casein or immunoglobulins or from the formation of lactoferrin multimers in the secretions. The interaction of lactoferrin with other proteins in mammary secretions during involution may affect the antimicrobial properties of lactoferrin.     

Lymphocyte phenotyping in infants: maturation of lymphocyte subpopulations and the effects of HIV infection

Oxygen administration is one of the most important therapeutic interventions for a child with severe acute lower respiratory tract infection (ALRI). Inexpensive and efficient methods of oxygen administration are highly desirable in hospitals in developing countries. The objectives of this study were to compare the frequency and nature of complications when nasopharyngeal catheters or nasal prongs are used to deliver oxygen. One hundred and twenty-one children between the ages of 2 weeks and 5 years with hypoxia due to ALRI were randomized to receive oxygen via a catheter (61 children) or via nasal prongs (60 children). The two groups were similar in terms of diagnoses, clinical severity, oxygen saturation on admission and case fatality rates. There was no difference in the incidence of hypoxaemic episodes between the two groups. The oxygen flow rates required on the day of admission for adequate oxygenation (SaO2 > 90%) ranged from 0.8 litres per minute to 1.2 litres per minute. The required oxygen flow rate decreased during the course of treatment. Mucus production was more of a problem in the catheter group, and nasal blockage, intolerance of the method of oxygen administration and nursing effort were generally higher amongst the catheter group, but none of these differences was significant. Ulceration or bleeding of the nose was significantly more common in the catheter group (19.7% vs 6.7%, p < 0.05). Abdominal distension and nasal perforation were not seen in either group. This study suggests that nasal prongs are safer, more comfortable and require less nursing expertise than nasopharyngeal catheters for administration of oxygen to children.     

Human gamma-interferon expression in the mammary gland of transgenic mice

Transgenic mice carrying a hybrid gene consisting of ovine beta-lactoglobulin gene sequences and human gamma-interferon (hIFN-g) cDNA were produced. hIFN-g expression in the mammary gland of two lactating transgenic founder females was found. The concentration of active hIFN-g in the milk was estimated as being ca. 1800 IU/ml. The hIFN-g ability to express in the mammary gland was found in the progeny of transgenic founder male.     

Environmental streptococcal intramammary infections of the bovine mammary gland

Characteristics of environmental streptococcal IMI were investigated over a 7-yr period for a herd in total confinement. A total of 374 new environmental streptococcal IMI was detected. Approximately 50.5% of IMI were new in the dry period, and 49.5% were new in lactation. The rate of new IMI was .00312 IMI/cow day during the dry period and .00054 IMI/cow day during lactation. The percentages of cows and quarters with an environmental streptococcal IMI present at calving were 10.6 and 3.2%, respectively. The percentage of heifers with an environmental streptococcal IMI at calving was similar to that for cows. The rate of new environmental streptococcal IMI was greater during the 1st mo of lactation than during the remainder of lactation. The rate of IMI during late lactation was higher for older cows than for either heifers or cows in second lactation. The rate of environmental streptococcal IMI during the dry period and during lactation was greatest during summer. The mean days of lactation that cows were infected for all IMI was 12.3 d. Approximately 41% of IMI had a duration of < 8 d. Stage of lactation, season of the year, and parity influenced the rate of new IMI.     

Telomerase activity in the normal and neoplastic rat mammary gland

The 1-methyl-1-nitrosourea-induced rat mammary tumor model system is well studied, reproducible, and widely used. We have investigated whether these tumors possess higher telomerase activity than normal mammary tissue. Using the telomeric repeat amplification protocol assay, we found significantly higher telomerase activity in 36 mammary carcinomas than in 72 mammary glands of virgin rats. The level of telomerase activity in virgin rats was unaffected by strain, age, stage of the estrous cycle, or ovariectomy. However, mammary glands obtained from pregnant rats exhibited telomerase activity comparable to that found in the tumors, possibly reflecting the high epithelial content of these tissues. Indeed, isolated epithelial cells from virgin and pregnant mammary glands and from carcinomas had similar telomerase activities. Thus, telomerase activity is constitutive in the rat mammary epithelium and is not a unique characteristic of malignant transformation in this tissue. These results underscore the importance of attributing biochemical properties to specific cell types in a tissue, a situation not paralleled in the interpretation of data from in vitro models.     

Contemporary methods of the target puncture of the mammary gland

New accurate methods of puncture of the mammary gland, making use of preliminary layout using roentgenograms, coordination nets, Cytoguide comprter device, and ultrasound-monitored spot biopsy replaced the traditional poorly informative methods. Sonography-monitored puncture is the most informative in cases with both palpated and unpalpated formations shaped as nodules: it permits imaging in 85% of cases. However, in 25-30% of cases unpalpated cancer presents as microcalcinates and cord-like restructuring which cannot be imaged on sonograms. Moreover, unpalpated cancer presenting as a nodule against the background of fatty involution also may be undiscernible on sonograms. In such cases puncture controlled by roentgenography with a computer device and a relevant kit of disposable instruments is advisable, giving accurate information in 61.1% of cases.     

Lymphocyte subpopulations of workers in a plant producing plastic materials (preliminary study)

Lymphocyte subpopulations were studied in 31 men working in a plant producing plastic materials in relation with control groups of similar age and smoking habit. 8 workers (group A) were exposed to solvents (mainly methylethylketone and dimethylformamide), 8 men (group B) to dust containing particles of calcium carbonate, polyvinylchloride, phtalates, unsaturated oils, paraffin wax, iron oxides, titanium bioxides, barium, zinc and lead and 15 men (group C), working in the same department as group B, were studied after a period of 16 months during which lead chromate was employed in the preparation of colors. The lymphocyte subpopulations were normal in group A, while in B there was a significant increase of HLA-DR + cells (monocytes, B and activated T lymphocytes). In group C, T helper/inducer lymphocytes (mainly CD4(+)-CD45RO- "virgin" lymphocytes), CD19+ B lymphocytes, CD3-HLADR+ and CD3-CD25+ (activated B lymphocytes and monocytes) were significantly reduced without changes of serum IgM, IgG and IgA. Highly significant correlation was found between B lymphocytes (reduced in the workers about 40%) and CD4(+)-CD45R0+ "memory" lymphocytes (reduced about 20%). Moreover, blood lead (correlated with urinary chromium) showed a highly significant negative correlation with the B lymphocytes. This study demonstrates that combined exposure to toxic agents produces specific modifications in the lymphocyte subsets without changes in immunoglobulins and confirms the results of previous researches showing that the exposure to lead or chromate induces reduction of lymphocytes in the peripheral blood.     

Osteopontin expression in mammary gland development and tumorigenesis

Osteopontin (OPN) is a secreted phosphoprotein that binds to cells via an Arg-Gly-Asp sequence and to mineralized surfaces. The protein can mediate cell adhesion and is strongly implicated in transformation and tumorigenesis. We have examined the expression pattern of OPN in mouse mammary glands at different stages of postnatal development. Whereas OPN is expressed at low-to-moderate levels in mammary glands from virgin and pregnant mice, the levels of OPN mRNA are extremely high in the lactating gland, consistent with the presence of the protein in milk. Expression is highest at 2 days of lactation and declines thereafter, but it remains high through involution. OPN expression is restricted to small nests or groups of cells at 9 days of involution. These results suggest that OPN may play a specific role in the process of involution that may be distinct from its role during lactation. In mammary tumors arising spontaneously in transgenic mice expressing the oncogenes c-myc and/or v-Ha-ras under the control of the mouse mammary tumor virus promoter, the level of OPN expression is increased dramatically over that in the normal gland in these same animals. Numerous cells expressing OPN mRNA are widespread throughout the tumors. OPN protein is detectable by Western blotting in extracts from the mammary gland at 2 days of lactation and from the tumors, but not in mammary glands at other stages of development. We hypothesize that OPN is exported from most tissues and that the protein is only detectable in tissues elaborating fluids, such as the lactating mammary gland, or in pathological situations when expression of OPN is abnormally high, such as in tumors.     

Expression of human lysozyme mRNA in the mammary gland of transgenic mice

Owing to its inherent antimicrobial effect and positive charge, the expression of human lysozyme in bovine milk could be beneficial by altering the overall microbial level and the functional and physical properties of the milk. We have used transgenic mice as model systems to evaluate the expression of human lysozyme containing fusion gene constructs in the mammary gland. Expression of human lysozyme was targeted to the mammary gland by using the 5' promoter elements of either the bovine beta (line B mice) or alpha s1 (line H mice) casein genes coupled to the cDNA for human lysozyme. Expression of human lysozyme mRNA was not found in mammary tissue from any of line B mice. Tissues were analysed from six lines of H mice and two, H6 and H5, were found to express human lysozyme mRNA in the mammary gland at 42% and 116%, respectively, of the levels of the endogenous mouse whey acidic protein gene. At peak lactation, female mice homozygous for the H5 and H6 transgene have approximately twice the amount of mRNA encoding human lysozyme as hemizygous animals. Expression levels of human lysozyme mRNA in the mammary gland at time points representing late pregnancy, early, peak and late lactation corresponded to the profile of casein gene expression. Human lysozyme mRNA expression was not observed in transgenic males, virgin females or in the kidney, liver, spleen or brain of lactating females. A very low level of expression of human lysozyme mRNA was observed in the salivary gland of line H5.     

Nerve and vessel supplying ligamentous suspension of the mammary gland

Anatomical findings from 28 breast specimens of female corpses have shown a thin horizontal fibrous septum, originating from the pectoral fascia along the level of the fifth rib, heading toward the nipple. This fibrous septum lies in between a cranial and a caudal vascular network, and being mesentery-like, it is responsible for the supply of the nipple areola complex. The cranial vascular sheet is supplied by the thoracoacromial artery and a branch of the lateral thoracic artery, whereas the caudal sheet is supplied by perforating branches from anastomoses of intercostal arteries. The fibrous septum is also a guiding structure for the main supplying nerve of the nipple. At its borders the septum curves upward into a vertical medial and lateral ligament, which attach the breast to the sternum and the lateral edge of pectoralis minor. These ligaments also contain a regular nerve and vascular supply. In their total, the fibrous septum and its ligaments form a sling of dense connective tissue that acts as a brassiere-like suspensory system. These two structures, the fibrous sling and the vascular and nervous membranes attached to it, are consistent anatomical findings, which have not been described before. Their knowledge could be of value and relevance in clinical application.     

Flow and composition of afferent mammary gland lymph

Afferent mammary lymphatic flow was characterized in conscious lactating cows during milking and prior to, during, and after intramammary infusion of endotoxins. Lymph flow (13 to 45 ml/h) was pulsatile with monophasic and multiphasic episodes. Flow resulted from 62 to 67 episodes per h. Episodes varied from 1 to 53 s in duration. Maximum instantaneous flow ranged from 163 to 245 microliters/s. Flow did not increase consistently during milking. Lymph flow increased (5.5- to 8-fold) during endotoxin-induced mastitis. Flow rates were elevated for up to 48 h after infusion of endotoxin. Compositional comparisons between afferent mammary lymph and blood plasma showed distinct differences. Lymph contained 7, 6, and 10 times less protein, albumin, and globulin, respectively, than did plasma. Glucose concentrations were equivalent. Lymph had 17 times less cholesterol and about one-half as much triglyceride and calcium as did plasma. gamma-Glutamyl-transaminase and aspartate transaminase were substantially higher in plasma than in lymph. Afferent mammary lymph has unique compositional characteristics. The lymph ducts contained an intrinsic mechanism for lymph movement. Moreover, this mechanism was altered by inflammation. The techniques herein provide a better understanding of the mammary lymphatic system.     

Intracellular behaviour of samarium and europium in lactating mammary gland

The subcellular localization of samarium and europium, two rare-earths, increasingly used in both medical and industrial fields, has been studied in several organs such as liver and kidney but never in the mammary gland despite of its importance in the biology of lactation and nutrition domains. The intracellular behaviour of samarium and europium after their intra-peritoneal administration in the lactating mammary gland cells was investigated. The results showed the presence of very electron dense deposits in the glandular epithelial cell lysosomes. These particular lysosomes were never observed in the mammary cell lysosomes of control rats. These intralysosomal deposits were probably composed of insoluble samarium or europium phosphates by analogy with previous studies, the transmission electron microscopy, the ion mass microscopy and the electron probe microanalysis, and other techniques allowing the identification of the chemical structure of the intralysosomal deposits.     

A paracrine role for the epithelial progesterone receptor in mammary gland development

Recently generated progesterone receptor (PR)-negative (PR-/-) mice provide an excellent model for dissecting the role of progesterone in the development of the mammary gland during puberty and pregnancy. However, the full extent of the mammary gland defect in these mice caused by the absence of the PR cannot be assessed, because PR-/- mice do not exhibit estrous cycles and fail to become pregnant. To circumvent this difficulty, we have transplanted PR-/- breasts into wild-type mice, and we have demonstrated that the development of the mammary gland in the absence of the PR is arrested at the stage of the simple ductal system found in the young virgin mouse. Mammary transplants lacking the PR in the stromal compartment give rise to normal alveolar growth, whereas transplants containing PR-/- epithelium conserve the abnormal phenotype. Chimeric epithelia in which PR-/- cells are in close vicinity to PR wild-type cells go through complete alveolar development to which the PR-/- cells contribute. Together, these results indicate that progesterone acts by a paracrine mechanism on a subset of mammary epithelial cells to allow for alveolar growth and that expression of the PR is not required in all the cells of the mammary epithelium in order for alveolar development to proceed normally.     

Peptides generated from milk proteins in the bovine mammary gland during involution

OBJECTIVE: To determine if the interaction between isoniazid and hydralazine, consisting of increased hypotension accompanied by bradycardia, occurs with other vasodilators. METHODS: Blood pressure and heart rate responses to a number of vasodilators were determined in rats under chloralose-urethane, pretreated or not with 250 mg/kg of isoniazid. The influence of this dose of isoniazid on GABA levels in the hypothalamus and pons-medulla was assessed in other groups of rats. RESULTS: Increased hypotension and bradycardia following i.p. isoniazid were observed with dipyridamole, prazosin, pinacidil and hydralazine given i.v. Bradycardia without increased hypotension appeared with papaverine and verapamil, while increased hypotension with unchanged heart rate was observed with minoxidil and captopril. Isoniazid decreased GABA in the hypothalamus and pons-medulla. CONCLUSIONS: At the high dose used, isoniazid interacts with various vasodilators, irrespective of their mechanism of action. The interaction could be due to the influence of the drug on GABA levels at cardiovascular regulatory sites.