Temporal fluctuations of cytokine concentrations in human milk

Circadian rhythms affect the circulating levels of proteins that can modulate immune responses. Cytokines represent a group of proteins present in breast milk that modulate the immune system, but the effects of fluctuations of these proteins have not yet been elucidated. In this work, the cytokines present in human milk were quantified, taking into account the phases of the day and the maturation stages of human milk. Samples were collected at different stages of milk maturation and in two phases, day and night. The levels of cytokines IL-2, IL-4, IL-6, IL-10, TNF-α, INF-γ and IL-17 were evaluated by flow cytometry. All quantified cytokines showed chronobiological variations with regard to the phase of day and/or stage of breast milk maturation. These data indicate that cytokines are subjected to fluctuations, and this knowledge is important for the use of human breast milk as an intervention strategy.     

Coenzyme Q concentration and total antioxidant capacity of human milk at different stages of lactation in mothers of preterm and full-term infants

Coenzyme Q₁₀(CoQ₁₀) in human milk at different stages of maturity in mothers of preterm and full-term infants and its relation to the total antioxidant capacity of milk is described for the first time. Thirty healthy breastfeeding women provided colostrum, transition-milk and mature-milk samples. Coenzyme Q, α-, γ- and δ-tocopherol, fatty acids and the total antioxidant capacity of the milk were analyzed. Coenzyme Q₁₀ was found at higher concentrations for colostrum (0.81 ± 0.06 vs. 0.50 ± 0.05 μmol/l) and transition milk (0.75 ± 0.06 vs. 0.45 ± 0.05 μmol/l) in the full-term vs. the preterm group (similar results were found for total antioxidant capacity). Concentrations of α- and γ-tocopherol were higher in the full-term group and decreased with time. In conclusion, CoQ₁₀ is present in breast milk, with higher concentration in mothers of full-term infants. CoQ₁₀ in breast milk decreases through lactation in mothers delivering full-term infants. Also, CoQ₁₀, α- and γ-tocopherol concentration in human milk directly correlates with the antioxidant capacity of the milk.     

母乳联合鼠李糖乳杆菌预防早产儿喂养不耐受的疗效观察

目的 观察母乳联合鼠李糖乳杆菌预防早产儿喂养不耐受的临床疗效。方法 选取2016年2月至2017年2月武汉大学人民医院收治的80例生后24 h内入住本院儿科新生儿病房的早产儿,随机分配成2组,其中研究组40例,对照组40例。研究组以新鲜母乳联合鼠李糖乳杆菌早期开奶喂养,对照组以早产儿配方奶早期开奶喂养,比较2组早产儿喂养不耐受症的发生率及其恢复出生体重时间、达全胃肠道喂养时间、体重增长速度和住院时间的差异。结果 研究组患儿喂养不耐受发生率低于对照组,研究组恢复出生体重时间、达全胃肠道喂养时间及住院时间均短于对照组（P<0.05）,研究组患儿体重增加速度快于对照组（P<0.05）。结论 早产儿早期喂养母乳联合鼠李糖乳杆菌能够较早建立全胃肠道营养,减少早产儿喂养不耐受症的发生率,促进患儿体重增长,缩短恢复出生体重时间及达全胃肠道喂养时间,降低住院天数。     

A pilot study of synbiotic supplementation on breast milk mineral concentrations and growth of exclusively breast fed infants

Despite the crucial role of breast milk mineral contents for health and growth of the infants, they decrease with the duration of lactation. So, this pilot study aimed to determine the effects of synbiotic supplementation on breast milk mineral composition and infants’ growth. In this pilot, randomized, double-blind, placebo-controlled trial, 57 lactating mothers were randomly divided into two groups to receive a daily supplement of synbiotic (n = 30) or a placebo (n = 27) for 30 days. Breast milk zinc, copper, Iron, magnesium and, calcium concentrations were determined by flame atomic absorption spectrometry. Weight for age Z-score (WAZ) and height for age Z-score (HAZ) were assessed for infants. Dietary intake was collected from lactating women using the 24-h recall method. Data analyses were carried out using nutritionist IV, Epi Info and SPSS soft wares. Synbiotic supplementation led to an insignificant increase of the mean breast milk levels of zinc (from 2.44 ± 0.65 to 2.55 ± 0.55 mg L⁻¹), copper (from 0.35 ± 0.24 to 0.40 ± 0.26 mg L⁻¹), iron (from 0.28 ± 0.42 to 0.31 ± 0.38 mg L⁻¹), magnesium (from 17.14 ± 1.35 to 17.17 ± 1.09 mg L⁻¹), and calcium (from 189 ± 25.3 to 189.9 ± 21.7 mg L⁻¹); whilst in the placebo group, these variables decreased significantly (P = 0.001). The observed changes between two groups were statistically significant (P < 0.05). Although WAZ and HAZ of infants increased slightly in the supplemented group (from 1.19 ± 0.79 to 1.20 ± 0.69 and 0.36 ± 0.86 to 0.37 ± 0.85 respectively), these two parameters decreased in the placebo group which was significant only for WAZ (P = 0.01). Moreover, no significant association was found between mineral intake and breast milk mineral contents. It seems, synbiotic supplementation may have positive effects on breast milk mineral contents.     

A comparative study of elemental composition of human breast milk and infant milk substitutes

Instrumental neutron activation analysis (INAA) and protoninduced X-ray emission (PIXE) analysis have been employed to determine the concentration of 13 elements in human breast milk, various infant formulas, and locally produced cereals from Nigeria, as well as from various infant formulas and natural cow and goat milk available in the UK. The study shows that if the locally produced cereal is to be used on a regular basis for babies in Nigeria, then their diet must be supplemented with essential trace elements. Furthermore, parents should be discouraged from giving their infants cow and goat milk because of the high concentration of major elements compared to human breast milk.     

Trace element content in human milk during lactation of preterm newborns

The present study has been finalized to perform the content of Zn, Cu, Cr, Se, Mn, F, Mo, Ni, and B in the preterm human milk over 21 d of lactation. Trace element (TE) contents were analyzed by inductively coupled plasma atomic emission spectroscopy (ICP-MS), and median concentrations of Zn, Cu, Cr, Se, Mn, and F observed in preterm milk did not demonstrate significant differences in comparison to levels shown in term milk. A statistical significant difference (p<0.05) has been found among Mo, Ni, and B content in preterm milk for every stage of lactation. TE content of infant blood founded concentrations of Mo in preterm babies significantly (p<0.01) lower than in term offsprings. Similar values of other TE content were obtained in blood of preterm, and term newborns. These findings point to the need for a considerable reassessment of the existing dietary recommendation for Mo content in infant feeding.     

Elemental composition of human milk from mothers of premature and full-term infants during the first 3 months of lactation

To examine longitudinal and gestational effects of mineral content in human milk, we analyzed human milk from lactating mothers of premature (PRT,n = 24, < 2000g birth weight, < 37 wk gestation) and fullterm (FT,n = 19, > 2500g, 39–41 wk gestation), living in Newfoundland, Canada. Samples were collected once a week for 8 wk with one final sample collected at 3 mo. Milk samples collected in acid-washed containers were wet ashed with concentrated HNO₃, and barium, cadmium, calcium, cesium, cobalt, copper, cerium, lanthanum, magnesium, manganese, molybdenum, nickel, lead, rubidium, tin, strontium, and zinc were measured using inductively coupled plasma-mass spectrometry. Data were analyzed using standard multiple-regression procedures with correlated data analyses to take account of the relationship between successive weeks. Results indicated lower Ca and Pb in PRT milk. Calcium was the only nutritionally significant element to differ between groups. Molybdenum in both PRT and FT milk showed a definite decrease with time, suggesting that the Mo content in milk is homeostatically regulated. However, Ce, La, Ba, and Sn did not display any pattern indicative of biological regulation and potential human requirement.     

Structural analysis of three sulfated oligosaccharides isolated from human milk

The structures of two sulfated octasaccharides and one sulfated nonasaccharide isolated from human milk have been investigated. Using 13C and 1H NMR spectroscopy and ESMS, the following structures 1-3 were established: [formula: see text].     

In vivo digestomics of milk proteins in human milk and infant formula using a suckling rat pup model

Human milk is the optimal mode of infant feeding for the first several months of life, and infant formulas serve as an alternative when breast-feeding is not possible. Milk proteins have a balanced amino acid composition and some of them provide beneficial bioactivities in their intact forms. They also encrypt a variety of bioactive peptides, possibly contributing to infant health and growth. However, there is limited knowledge of how milk proteins are digested in the gastrointestinal tract and bioactive peptides are released in infants. A peptidomic analysis was conducted to identify peptides released from milk proteins in human milk and infant formula, using a suckling rat pup model. Among the major milk proteins targeted, α-lactalbumin and β-casein in human milk, and β-lactoglobulin and β-casein in infant formula were the main sources of peptides, and these peptides covered large parts of the parental proteins’ sequences. Release of peptides was concentrated to specific regions, such as residues 70–92 of β-casein in human milk, residues 39–55 of β-lactoglobulin in infant formula, and residues 57–96 and 145–161 of β-CN in infant formula, where resistance to gastrointestinal digestion was suggested. In the context of bioactive peptides, release of fragments containing known bioactive peptides was confirmed, such as β-CN-derived opioid and antihypertensive peptides. It is therefore likely that these fragments are of biological significance in neonatal health and development.     

Peptidome analysis of human skim milk in term and preterm milk

The abundant proteins in human milk have been well characterized and are known to provide nutritional, protective, and developmental advantages to both term and preterm infants. Due to the difficulties associated with detection technology of the peptides, the expression of the peptides present in human milk is not known widely. In recent years, peptidome analysis has received increasing attention. In this report, the analysis of endogenous peptides in human milk was done by mass spectrometry. A method was also developed by our researchers, which can be used in the extraction of peptide from human milk. Analysis of the extracts by LC–MS/MS resulted in the detection of 1000–3000 Da peptide-like features. Out of these, 419 peptides were identified by MS/MS. The identified peptides were found to originate from 34 proteins, of which several have been reported. Analysis of the peptides’ cleavage sites showed that the peptides are cleaved with regulations. This may reflect the protease activity and distribution in human body, and also represent the biological state of the tissue and provide a fresh source for biomarker discovery. Isotope dimethyl labeling analysis was also used to test the effects of premature delivery on milk protein composition in this study. Differences in peptides expression between breast milk in term milk (38–41 weeks gestation) and preterm milk (28–32 weeks gestation) were investigated in this study. 41 Peptides in these two groups were found expressed differently. 23 Peptides were present at higher levels in preterm milk, and 18 were present at higher levels in term milk.     

A physiological role of breast milk leptin in body weight control in developing infants

Objective: Leptin, a hormone that regulates food intake and energy metabolism, is present in breast milk. The aim of this study was to determine whether milk leptin concentration is correlated with maternal circulating leptin and BMI and with body weight gain of infants. Research Methods and Procedures: A group of 28 non‐obese women (BMI between 16.3 and 27.3 kg/m²) who breast‐fed their infants for at least 6 months and their infants were studied. Venous blood and milk samples were obtained from mothers at 1, 3, 6, and 9 months of lactation, and leptin concentration was determined. Infant body weight and height were followed until 2 years of age. Results: During the whole lactation period, milk leptin concentration correlated positively with maternal plasma leptin concentration and with maternal BMI. In addition, milk leptin concentration at 1 month of lactation was negatively correlated with infant BMI at 18 and 24 months of age. A better negative correlation was also found between log milk leptin concentration at 1 and at 3 months of lactation and infant BMI from 12 to 24 months of age. Discussion: We concluded that, in a group of non‐obese mothers, infant body weight during the first 2 years may be influenced by milk leptin concentration during the first stages of lactation. Thus, moderate milk‐borne maternal leptin appears to provide moderate protection to infants from an excess of weight gain. These results seem to point out that milk leptin is an important factor that could explain, at least partially, the major risk of obesity of formula‐fed infants with respect to breast‐fed infants.     

原料乳冷藏过程中微生物细胞的宏蛋白组学分析

原料乳在投入生产前通常需4°C冷藏,在这个过程中微生物的污染将造成冷藏原料乳的变质。【目的】本文旨在分子水平上探究原料乳冷藏过程中微生物表达的差异蛋白质的动态变化,为原料乳的冷藏提供理论支撑。【方法】利用Label-free技术研究原料乳4°C冷藏6 d期间微生物菌体蛋白质的物种来源、功能及参与通路,筛选差异蛋白质并对主要差异蛋白质参与的通路进行富集,探究其变化规律。【结果】冷藏过程中共鉴定出341个微生物菌体蛋白质,其中冷藏3 d后产生的蛋白质占所有检出蛋白质的60.12%。COG及KEGG分析表明,蛋白质所体现的功能及参与的通路随时间而变化。冷藏4 d,参与糖酵解/糖异生、ABC转运蛋白、氨基糖和核苷酸糖代谢等通路的蛋白数量显著增加。随冷藏时间延长,相邻时间点差异蛋白质数目逐渐增多,且富集在不同的通路中。【结论】冷藏过程中原料乳中的微生物所产生的蛋白质参与的通路及其所体现的功能复杂,4 d时的变化最为明显,或可作为原料乳质量控制的关键节点。     

Alterations in the milk yield and composition during different stages of lactation cycle in elite and non-elite Karan-Fries cross-bred cows (Holstein Fresian x Tharparkar)

Alterations in the milk constituents throughout the lactation cycle have been studied in 20 Karan-Fries cross-bred cows (Holstein Fresian x Tharparkar) divided into two groups, viz. elite (n = 10) and non-elite (n = 10). About 100 ml of composite milk samples (from all the quarters) were collected through hand milking in sterile tubes from the day of calving at 15-day interval till 300 days of lactation cycle. Different milk constituents (viz. fat, protein, lactose, Solid not fat) were estimated by automatic milk analyzer. The daily milk yield varied significantly (p < 0.01) between group and different days of lactation cycle. Milk fat percentage of milk did not differ significantly between groups and different days of lactation cycle. Milk protein and lactose percentage did not differ significantly between groups but differed significantly (p < 0.001) between days. Solid not fat (SNF) percentage of milk was significantly (p < 0.01) higher in elite cows compared to non-elite cows and also varied significantly (p < 0.05) during different days of lactation cycle. In both the group of cows, milk protein, fat, and SNF percentage was highest and lactose percentage was lowest up to 1st week of lactation cycle and rapidly declined thereafter due to the transition of the colostrum into milk. After 2nd week of lactation, all the milk constituents under study were almost remained unaltered till the end of the cycle. The above stated investigation not only substantiates the already existing information of lactation stage-specific alteration in milk constituents but also depict the exact point of transition of these constituents during the lactation cycle so the nutritional and managemental interventions could be carried out in proper time.     

母乳联合妈咪爱对极低出生体重儿炎性细胞因子的影响

目的 探讨母乳联合妈咪爱对极低出生体重儿（ELBWI）炎性细胞因子的影响。方法 采用前瞻性研究法,选择2012年2月至2016年5月在我院诊治的ELBWI 76例,根据随机数字表法分为观察组与对照组各38例,对照组给予母乳喂养,观察组给予母乳联合妈咪爱喂养,两组均给予暖箱保暖、治疗原发病等对症治疗。治疗14 d后,观察两组患儿的预后与炎性细胞因子表达情况。结果 观察组与对照组的喂养不耐受率分别为7.9%和26.3%,观察组明显少于对照组（χ2=3.289,P0.05）。观察组治疗后的血清IL-6与TNF-α含量分别为（16.33±11.61）ng/L和（25.23±8.92）ng/L,均明显低于对照组的（26.35±11.56）ng/L与（33.72±10.36）ng/L（P<0.05）,且都明显低于治疗前（P<0.05）。结论 母乳联合妈咪爱喂养可改善ELBWI早期喂养的耐受性,提高患儿免疫功能,降低炎性细胞因子IL-6与TNF-α的表达,从而对生长发育有一定促进作用。     

Alteration in the in vitro activity of milk leukocytes during different parity in high yielding cross-bred cows

In vitro activity of milk leukocytes (viz. neutrophils, lymphocytes and macrophages) was evaluated in forty-eight (48) clinically healthy high-yielding cross-bred cows of mid-lactation stage (100–200 days of lactation), divided into four groups namely 1st parity (n = 12), 2nd parity (n = 12), 3rd parity (n = 12) and 4th and above parity (n = 12). Milk samples were taken (250 ml/cow) were taken. Milk somatic cell counts (SCC) and differential leukocyte counts (DLC) were performed microscopically. In vitro phagocytic index (PI) of milk neutrophils and macrophages was evaluated by colorimetric nitro blue tetrazolium reductive assay. Mitogen-induced milk lymphocyte blastogenic response was measured by colorimetric MTT (tetrazolium) assay after isolation of the milk leukocytes by density gradient centrifugation. Milk SCC differed significantly (p < 0.01) between different parity. Cows of 4 and above parity showed significantly (p < 0.01) higher milk SCC compared to primiparous cows. There was no significant difference in milk DLC during different parities in high-yielding cross-bred cows. There was a significant (p < 0.01) variation in lymphocyte blastogenesis amongst parity. The highest value of lymphocyte blastogenesis was seen at 3rd parity, whereas lowest value was obtained in the cows of both 1st and 4th or above parity. PI of milk neutrophils did not differ significantly between parity. PI of milk macrophages was significantly (p < 0.01) higher in 3rd parity and lower (p < 0.01) in 1st and 4th parities. The study indicated that depressed activity of milk lymphocytes and macropages was lower and SCC was higher in the cows of 4th and above parity indicating more mammary stress and hence susceptible to udder infection and mastitis. Therefore, better care and managemental interventions should be taken around these periods.     

Enkephalin binding systems in human plasma III: Comparative protection of different peptides

We have investigated the hydrolysis and protection from hydrolysis of several peptides by plasma enzymes and by the plasma components previously described as inhibitors of enkephalins' hydrolysis. The results shown indicate that all the peptides actually hydrolyzed are also partially protected from hydrolysis by the enkephalin-protecting substances. Protection is fairly uniform for all the peptides tested, but considerably higher in the case of leu- and met-enkephalin, suggesting a partial specificity of the protecting substances towards opioid peptides.     

成都地区母乳双歧杆菌对婴儿肠道双歧杆菌构建的影响

观察不同喂养方式婴儿肠道双歧杆菌的构建规律,初步探索母乳双歧杆菌与婴儿肠道双歧杆菌之间的相关性。

在四川大学华西第二医院纳入57对母婴志愿者,根据喂养方式分为母乳（BF）组（n = 31）和混合喂养（MF）组（1月龄内添加奶粉时间＞8 d,n = 26）,收集产妇产后0天、第30天的母乳,以及婴儿出生后0天、第7天、第30天的粪便。采用实时荧光定量PCR测定母乳和婴儿粪便中双歧杆菌属、角双歧杆菌（B. angulatum）、链双歧杆菌（B. catnulatum）、齿双歧杆菌（B. dentium）、短双歧杆菌（B. breve）、两歧双歧杆菌（B. bifidum）、长双歧杆菌长亚种（B. longum subsp. longum）、长双歧杆菌婴儿亚种（B. longum subsp. infantis）及青春双歧杆菌（B. adolescentis）的丰度。观察2组婴儿肠道双歧杆菌0―7―30天的构建规律;分析母乳双歧杆菌与婴儿肠道双歧杆菌之间的相关性。

0―7―30天2组婴儿肠道双歧杆菌构建过程存在差异。母乳双歧杆菌与婴儿肠道双歧杆菌存在相关：0 d时,BF组母乳B. angulatum与婴儿肠道B. longum subsp. longum呈正相关（r = 0.962 3,P = 0.037 7）,母乳B. longum subsp. infantis与婴儿肠道B. longum subsp. longum、B. breve及B. dentium呈正相关（r = 0.998 4,P = 0.036 1;r = 1.000 0,P = 0.000 2;r = 1.000 0,P = 0.002 3）,母乳B. catnulatum与婴儿肠道B. dentium呈正相关（r = 0.997 7,P = 0.043 1）;30 d时,BF组母乳B. adolescentis与婴儿肠道双歧杆菌属呈正相关（r = 0.532 2,P = 0.013 0）,母乳B. longum subsp. longum与婴儿肠道B. breve、B. catnulatum呈负相关（r = ‒0.458 9,P = 0.048 1;r = ‒0.520 7,P = 0.032 1）,母乳B. catnulatum与婴儿肠道B. angulatum呈负相关（r = ‒0.682 8,P = 0.007 1）,母乳双歧杆菌属与婴儿肠道B. longum subsp. longum呈负相关（r = ‒0.714 3,P = 0.046 5）;30 d时,MF组母乳B. bifidum与婴儿肠道B. longum subsp. longum呈正相关（r = 0.579 2,P = 0.030 0）,母乳B. angulatum与婴儿肠道B. catnulatum呈正相关（r = 0.550 1,P = 0.022 1）。

喂养方式会影响婴儿肠道双歧杆菌构建;母乳中存在双歧杆菌且与婴儿肠道双歧杆菌相关,母乳喂养与婴儿肠道双歧杆菌的相关性更密切,尤其0 d时相关性更强;混合喂养可能由于奶粉的添加使母乳双歧杆菌与婴儿肠道双歧杆菌的相关性降低,规律性减弱。

     

Characterization of gut microbiota and short-chain fatty acid in breastfed infants with or without breast milk jaundice

This study aims to investigate the gut microbiota and metabolites in breastfed infants with breast milk jaundice (BMJ) using gut microbiome–metabolomics. Breastfed newborns diagnosed with BMJ and those without BMJ (control group) were enrolled. Faecal samples were collected from the participants and subjected to high-throughput sequencing of the 16s rDNA V3 and V4 regions of the gut flora and metabolomics of short-chain fatty acids (SCFAs). Proteobacteria, Fimicutes and Actinobacteria were the main bacteria at the phylum level. Eshcerichia-Shigella and Enterobacteriacea were the main bacteria at the genus level. The difference between the two groups was compared. Compared to the control group, the amount of Streptococcus was significantly increased while the amount of Enterococcus was significantly decreased in the faeces from infants with BMJ. Functional prediction analysis of 16S found that biosynthesis of penicillin and cephalosporin significantly increased in the BMJ group. Gas chromatography–mass spectrometry detection of SCFAs revealed that levels of acetic acid and propionic acid were significantly lower in the BMJ group than in the control group. The reduced levels of acetic acid and propionic acid may be related to the increase in Streptococcus and decrease in Enterococcus, both of which may contribute to BMJ.     

Growth of bifidobacteria and clostridia on human and cow milk saccharides

For healthy infants, which were born normally and fully breastfed, the dominant component of the intestinal microflora are bifidobacteria. However, infants born by caesarean section possess clostridia as a dominant intestinal bacterial group. The aim of the present study was to determine whether bifidobacteria and clostridia are able to grow on human milk oligosaccharides (HMOs) and other carbon sources - lactose, cow milk (CM) and human milk (HM). Both bifidobacteria and clostridia grew on lactose and in CM. Bifidobacteria grew in HM and on HMOs. In contrast, 3 out of 5 strains of clostridia were not able to grow in HM. No clostridial strain was able to utilise HMOs. While both bifidobacterial strains were resistant to lysozyme, 4 out of 5 strains of clostridia were lysozyme-susceptible. It seems that HMOs together with lysozyme may act as prebiotic-bifidogenic compounds inhibiting intestinal clostridia.     

Coenzyme Q concentration and total antioxidant capacity of human milk at different stages of lactation in mothers of preterm and full-term infants

Coenzyme Q10(CoQ10) in human milk at different stages of maturity in mothers of preterm and full-term infants and its relation to the total antioxidant capacity of milk is described for the first time. Thirty healthy breastfeeding women provided colostrum, transition-milk and mature-milk samples. Coenzyme Q, alpha-, gamma- and delta-tocopherol, fatty acids and the total antioxidant capacity of the milk were analyzed. Coenzyme Q10 was found at higher concentrations for colostrum (0.81+/-0.06 vs. 0.50+/-0.05 micromol/l) and transition milk (0.75+/-0.06 vs. 0.45+/-0.05 micromol/l) in the full-term vs. the preterm group (similar results were found for total antioxidant capacity). Concentrations of alpha- and gamma-tocopherol were higher in the full-term group and decreased with time. In conclusion, CoQ10 is present in breast milk, with higher concentration in mothers of full-term infants. CoQ10 in breast milk decreases through lactation in mothers delivering full-term infants. Also, CoQ10, alpha- and gamma-tocopherol concentration in human milk directly correlates with the antioxidant capacity of the milk.