Publications by authors named "Shannon L Kelleher"

85 Publications

Macrolets: Outsized Extracellular Vesicles Released from Lipopolysaccharide-Stimulated Macrophages that Trap and Kill Escherichia coli.

iScience 2020 Jun 5;23(6):101135. Epub 2020 May 5.

Department of Surgery, Penn State College of Medicine and Milton S. Hershey Medical Center, Room# C4810, H149, 500 University Drive, Hershey, PA 17033, USA; Department of Cellular & Molecular Physiology, Penn State Hershey College of Medicine, Hershey, PA 17033, USA. Electronic address:

Macrophages release a variety of extracellular vesicles (EVs). Here we describe a previously unreported class of EVs that are released from macrophages in response to Escherichia coli endotoxin, lipopolysaccharide (LPS), that we have named "macrolets" since they are extruded as large "droplets" released from macrophages. Morphologically, macrolets are anuclear, bounded by a single lipid membrane and structurally dependent on an actin cytoskeleton. Macrolets are enriched in tetraspanins and separable on this basis from their parent macrophages. Macrolets are distinguished from classic exosomes by their larger size (10-30 μm), discoid shape, and the presence of organelles. Macrolets are rich in both interleukin 6 (IL-6) and interleukin 6 receptor (IL-6R),and are capable of trapping and killing E. coli in association with production of reactive oxygen species. Our observations offer insights into the mechanisms by which macrophage activities may be amplified in sites of infection, inflammation, and healing.
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http://dx.doi.org/10.1016/j.isci.2020.101135DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7240733PMC
June 2020

A common genetic variant in zinc transporter ZnT2 (Thr288Ser) is present in women with low milk volume and alters lysosome function and cell energetics.

Am J Physiol Cell Physiol 2020 06 22;318(6):C1166-C1177. Epub 2020 Apr 22.

Department of Biomedical and Nutritional Sciences, University of Massachusetts Lowell, Lowell, Massachusetts.

Suboptimal lactation is a common, yet underappreciated cause for early cessation of breastfeeding. Molecular regulation of mammary gland function is critical to the process lactation; however, physiological factors underlying insufficient milk production are poorly understood. The zinc (Zn) transporter ZnT2 is critical for regulation of mammary gland development and maturation during puberty, lactation, and postlactation gland remodeling. Numerous genetic variants in the gene encoding ZnT2 () are associated with low milk Zn concentration and result in severe Zn deficiency in exclusively breastfed infants. However, the functional impacts of genetic variation in ZnT2 on key mammary epithelial cell functions have not yet been systematically explored at the cellular level. Here we determined a common mutation in /ZnT2 substituting serine for threonine at amino acid 288 (Thr288Ser) was found in 20% of women producing low milk volume ( = 2/10) but was not identified in women producing normal volume. Exploration of cellular consequences in vitro using phosphomimetics showed the serine substitution promoted preferential phosphorylation of ZnT2, driving localization to the lysosome and increasing lysosome biogenesis and acidification. While the substitution did not initiate lysosome-mediated cell death, cellular ATP levels were significantly reduced. Our findings demonstrate the Thr288Ser mutation in /ZnT2 impairs critical functions of mammary epithelial cells and suggest a role for genetic variation in the regulation of milk production and lactation performance.
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http://dx.doi.org/10.1152/ajpcell.00383.2019DOI Listing
June 2020

Milk-derived miRNA profiles elucidate molecular pathways that underlie breast dysfunction in women with common genetic variants in SLC30A2.

Sci Rep 2019 09 3;9(1):12686. Epub 2019 Sep 3.

Department of Surgery, Penn State Hershey College of Medicine, Hershey, Pennsylvania, United States of America.

Studies in humans and pre-clinical animal models show milk-derived miRNAs reflect mammary gland function during lactation. The zinc transporter SLC30A2/ZnT2 plays a critical role in mammary gland function; ZnT2-null mice have profound defects in mammary epithelial cell (MEC) polarity and secretion, resulting in sub-optimal lactation. Non-synonymous genetic variation in SLC30A2 is common in humans, and several common ZnT2 variants are associated with changes in milk components that suggest breast dysfunction in women. To identify novel mechanisms through which dysfunction might occur, milk-derived miRNA profiles were characterized in women harboring three common genetic variants in SLC30A2 (DE, TS, and Exon 7). Expression of ten miRNAs differed between genotypes, and contributed to distinct spatial separation. Studies in breast milk and cultured MECs confirmed expression of ZnT2 variants alters abundance of protein levels of several predicted mRNA targets critical for breast function (PRLR, VAMP7, and SOX4). Moreover, bioinformatic analysis identified two novel gene networks that may underlie normal MEC function. Thus, we propose that genetic variation in genes critical for normal breast function such as SLC30A2 has important implications for lactation performance in women, and that milk-derived miRNAs can be used to identify novel mechanisms and for diagnostic potential.
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http://dx.doi.org/10.1038/s41598-019-48987-4DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6722070PMC
September 2019

The Role of Genetic Variant rs13266634 in SLC30A8/ZnT8 in Post-Operative Hyperglycemia after Major Abdominal Surgery.

J Clin Endocrinol Metab 2019 Apr 10. Epub 2019 Apr 10.

Department of Surgery, Penn State Hershey College of Medicine and Milton S. Hershey Medical Center, Hershey, PA.

Context: Following major surgery, post-operative hyperglycemia (POHG) is associated with suboptimal outcomes, among diabetics and non-diabetics. A specific genetic variant, rs13266634 (c.973C>T; p.ARG325TRP) in zinc transporter SLC30A8/ZnT8, is associated with protection against Type-2 Diabetes, suggesting it may be actionable for predicting and preventing POHG.

Objective: To determine independent and mediated influences of a genetic variant on POHG in patients undergoing a model major operation, complex abdominal ventral hernia repair (cVHR).

Patients And Methods: For 110 patients (mean BMI 34.9±5.8, T2D history 28%) undergoing cVHR at a tertiary referral center (January 2012 to March 2017), multivariate regression was used to correlate the rs13266634 variant to pre-operative clinical, laboratory and imaging-based indices of liver steatosis and central abdominal adiposity to POHG. Causal Mediation Analysis (CMA) was used to determine direct and mediated contributions of SLC30A8/ZnT8 status to POHG.

Results: Variant rs13266634 was present in 61 patients (55.4%). In univariate models, when compared to patients with rs13266634, the homozygous wild-genotype (C/C, n=49) was associated with significantly higher risks of POHG (OR= 0.30 95%CI =0.14, 0.67, P=0.0038). Multivariate regression indicated that the association was independent (OR= 0.39 95%CI 0.15-0.97, p=0.040). In addition, CMA suggested that rs13266634 protects against POHG directly and indirectly through its influence on liver steatosis and central adiposity.

Conclusions: In medically complex patients undergoing major operations, the rs13266634 variant protects against POHG and its associated outcomes, through independent and mediated contributions. In C/C patients undergoing major operations, SLC30A8/ZnT8 may prove useful to stratify risk of POHG and potentially as a therapeutic target.
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http://dx.doi.org/10.1210/jc.2018-02588DOI Listing
April 2019

Increased early enteral zinc intake improves weight gain in hospitalised preterm infants.

Acta Paediatr 2019 11 27;108(11):1978-1984. Epub 2019 May 27.

Division of Neonatal-Perinatal Medicine, Penn State Health Children's Hospital, Hershey, PA, USA.

Aim: To test the hypothesis that enteral zinc intake is associated with improved preterm infant growth during neonatal intensive care unit (NICU) hospitalisation.

Methods: This prospective cohort study enrolled 105 preterm infants at a tertiary referral centre. Enteral zinc intake was calculated at day of life 14, and growth was measured as change in weight, length and head circumference from birth to discharge. Nonparametric tests assessed the contribution of breast milk vs formula and enteral zinc intake on weight, length and head circumference growth. Partial correlations evaluated the impact of baseline health status and caloric intake on growth. Multiple regression analysis was then completed to determine the unique contribution of zinc intake to weight gain and head circumference growth.

Results: Total enteral zinc intake was positively associated with weight gain (r = 0.4, p < 0.01) and head circumference growth (r = 0.3, p < 0.01) during NICU hospitalisation. Further, multiple regression analysis showed higher zinc intake is linked to weight gain during NICU hospitalisation after accounting for postmenstrual age at birth.

Conclusion: Increased early enteral zinc intake is linked to weight gain during NICU hospitalisation, highlighting the importance of enteral zinc intake in early infant nutrition.
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http://dx.doi.org/10.1111/apa.14828DOI Listing
November 2019

Excess Dietary Zinc Intake in Neonatal Mice Causes Oxidative Stress and Alters Intestinal Host-Microbe Interactions.

Mol Nutr Food Res 2019 02 12;63(3):e1800947. Epub 2018 Dec 12.

Department of Surgery, Penn State Hershey College of Medicine, Hershey, PA.

Scope: Greater than 68% of young infants are exposed to dietary zinc (Zn) levels that are higher than the Tolerable Upper Intake Limit. However, the consequences of excess dietary Zn during early life on intestinal function and host-microbe interactions are unknown.

Methods And Results: Neonatal mice are gavaged with 100 Zn µg d from postnatal day (PN) 2 through PN10 and indices of intestinal function and host-microbe interactions are compared to unsupplemented mice. Excess dietary Zn causes oxidative stress, increases goblet cell number and mucus production, and are associated with increased intestinal permeability and systemic inflammation. Over 900 genes are differentially expressed; 413 genes display a fold-change >1.60. The Gene Ontology Biological processes most significantly affected include biological adhesion, the immune system, metabolic processes, and response to stimulus. Key genes most highly and significantly upregulated include ALDH2, MT1, TMEM6, CDK20, and COX62b, while CALU, ST3GAL4, CRTC2, SLC28A2, and COMMA1 are downregulated. These changes are associated with a microbiome enriched in pathogenic taxa including Pseudomonadales and Campylobacter, and greater expression of bacterial stress response genes.

Conclusion: Excess dietary Zn may have unforeseen influences on epithelial signaling pathways, barrier function, and luminal ecology in the intestine that may have long-term consequences on intestinal health.
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http://dx.doi.org/10.1002/mnfr.201800947DOI Listing
February 2019

ZnT2 is critical for lysosome acidification and biogenesis during mammary gland involution.

Am J Physiol Regul Integr Comp Physiol 2018 08 2;315(2):R323-R335. Epub 2018 May 2.

Department of Cellular and Molecular Physiology, Penn State Hershey College of Medicine , Hershey, Pennsylvania.

Mammary gland involution, a tightly regulated process of tissue remodeling by which a lactating mammary gland reverts to the prepregnant state, is characterized by the most profound example of regulated epithelial cell death in normal tissue. Defects in the execution of involution are associated with lactation failure and breast cancer. Initiation of mammary gland involution requires upregulation of lysosome biogenesis and acidification to activate lysosome-mediated cell death; however, specific mediators of this initial phase of involution are not well described. Zinc transporter 2 [ZnT2 ( SLC30A2)] has been implicated in lysosome biogenesis and lysosome-mediated cell death during involution; however, the direct role of ZnT2 in this process has not been elucidated. Here we showed that ZnT2-null mice had impaired alveolar regression and reduced activation of the involution marker phosphorylated Stat3, indicating insufficient initiation of mammary gland remodeling during involution. Moreover, we found that the loss of ZnT2 inhibited assembly of the proton transporter vacuolar ATPase on lysosomes, thereby decreasing lysosome abundance and size. Studies in cultured mammary epithelial cells revealed that while the involution signal TNFα promoted lysosome biogenesis and acidification, attenuation of ZnT2 impaired the lysosome response to this involution signal, which was not a consequence of cytoplasmic Zn accumulation. Our findings establish ZnT2 as a novel regulator of vacuolar ATPase assembly, driving lysosome biogenesis, acidification, and tissue remodeling during the initiation of mammary gland involution.
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http://dx.doi.org/10.1152/ajpregu.00444.2017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7199225PMC
August 2018

A genetic variant in SLC30A2 causes breast dysfunction during lactation by inducing ER stress, oxidative stress and epithelial barrier defects.

Sci Rep 2018 02 23;8(1):3542. Epub 2018 Feb 23.

Department of Cellular and Molecular Physiology, Penn State Hershey College of Medicine, Hershey, PA, 17033, USA.

SLC30A2 encodes a zinc (Zn) transporter (ZnT2) that imports Zn into vesicles in highly-specialized secretory cells. Numerous mutations and non-synonymous variants in ZnT2 have been reported in humans and in breastfeeding women; ZnT2 variants are associated with abnormally low milk Zn levels and can lead to severe infantile Zn deficiency. However, ZnT2-null mice have profound defects in mammary epithelial cell (MEC) polarity and vesicle secretion, indicating that normal ZnT2 function is critical for MEC function. Here we report that women who harbor a common ZnT2 variant (TS) present with elevated levels of several oxidative and endoplasmic reticulum (ER) stress markers in their breast milk. Functional studies in vitro suggest that substitution of threonine for serine at amino acid 288 leads to hyperphosphorylation retaining ZnT2 in the ER and lysosomes, increasing ER and lysosomal Zn accumulation, ER stress, the generation of reactive oxygen species, and STAT3 activation. These changes were associated with decreased abundance of zona occludens-1 and increased tight junction permeability. This study confirms that ZnT2 is important for normal breast function in women during lactation, and suggests that women who harbor defective variants in ZnT2 may be at-risk for poor lactation performance.
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http://dx.doi.org/10.1038/s41598-018-21505-8DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5824919PMC
February 2018

Zinc transporter 2 interacts with vacuolar ATPase and is required for polarization, vesicle acidification, and secretion in mammary epithelial cells.

J Biol Chem 2017 12 7;292(52):21598-21613. Epub 2017 Nov 7.

From the Departments of Cellular and Molecular Physiology,

An important feature of the mammary gland is its ability to undergo profound morphological, physiological, and intracellular changes to establish and maintain secretory function. During this process, key polarity proteins and receptors are recruited to the surface of mammary epithelial cells (MECs), and the vesicle transport system develops and matures. However, the intracellular mechanisms responsible for the development of secretory function in these cells are unclear. The vesicular zinc (Zn) transporter ZnT2 is critical for appropriate mammary gland architecture, and ZnT2 deletion is associated with cytoplasmic Zn accumulation, loss of secretory function and lactation failure. The underlying mechanisms are important to understand as numerous mutations and non-synonymous genetic variation in ZnT2 have been detected in women that result in severe Zn deficiency in exclusively breastfed infants. Here we found that ZnT2 deletion in lactating mice and cultured MECs resulted in Zn-mediated degradation of phosphatase and tensin homolog (PTEN), which impaired intercellular junction formation, prolactin receptor trafficking, and alveolar lumen development. Moreover, ZnT2 directly interacted with vacuolar H-ATPase (V-ATPase), and ZnT2 deletion impaired vesicle biogenesis, acidification, trafficking, and secretion. In summary, our findings indicate that ZnT2 and V-ATPase interact and that this interaction critically mediates polarity establishment, alveolar development, and secretory function in the lactating mammary gland. Our observations implicate disruption in ZnT2 function as a modifier of secretory capacity and lactation performance.
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http://dx.doi.org/10.1074/jbc.M117.794461DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5766956PMC
December 2017

Peripheral zinc and neopterin concentrations are associated with mood severity in bipolar disorder in a gender-specific manner.

Psychiatry Res 2017 09 17;255:52-58. Epub 2017 May 17.

Department of Psychiatry, Penn State College of Medicine and Penn State Milton S. Hershey Medical Center, Hershey, PA 17033, USA; University of Michigan Department of Psychiatry and Depression Center, Ann Arbor, MI 48109, USA.

Bipolar disorder (BD) is a recurrent, episodic mood disorder for which there are no current diagnostic, prognostic or theranostic biomarkers. Two peripheral markers of the acute phase immune response, zinc and neopterin, are consistently associated with severity of depression in literature. Given gender differences in clinical presentation of BD and in inflammatory processes, we aimed to explore the interaction between gender and immune biomarkers to predict mood severity in BD. Participants with DSM IV BD I and II were recruited through the Pennsylvania Psychiatric Institute during an acute mood episode. Healthy controls (HC) were recruited through advertisements. Participants fasted for at least 6h when blood was drawn for biomarkers. We found that zinc concentrations were significantly lower in the BD group at baseline (p<.05), and there was also a significant interaction between gender and zinc (p<.05), associated with depression severity. Also, we found a significant interaction between gender and neopterin, associated with mania severity (p<.05). We found that mania severity was associated with neopterin in men, while depression severity was positively associated with zinc in women. Our report bears replication in larger samples and highlights the potential for differences in the underlying pathophysiology between men and women with BD.
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http://dx.doi.org/10.1016/j.psychres.2017.05.022DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5545151PMC
September 2017

Immune and metabolic responses in early and late sepsis during mild dietary zinc restriction.

J Surg Res 2017 04 2;210:47-58. Epub 2016 Nov 2.

Department of Surgery, Pennsylvania State University College of Medicine, Hershey, Pennsylvania; Department of Cellular and Molecular Physiology, Pennsylvania State University College of Medicine, Hershey, Pennsylvania. Electronic address:

Background: Mild dietary zinc (Zn) deficiency is widespread in human populations, but its influence on recovery after acute illness is poorly understood. In a mouse model of abdominal sepsis (cecal ligation puncture), systemic immune responses and liver metabolism were monitored in early (24 h) and late (5 d) phases, under control conditions and during mild dietary Zn restriction.

Methods: Mice were fed diets adequate or marginally deficient (ZM) in Zn (30 versus 10 mg zinc/kg diet) for 4 wk, before undergoing laparotomy alone (nonseptic control) or cecal ligation puncture (septic).

Results: Among nonseptic mice, the ZM state was not associated with differences in inflammation or metabolic responses. Among septic mice, mortality did not differ between the zinc adequate and ZM groups. In the early phase, the ZM state amplified increases in plasma interleukin (IL) 6, tumor necrosis factor alpha, and IL-10, while dampening the interferon gamma response. In the late phase, subtle but significant ZM-associated increases were observed in plasma IL-5 and interferon gamma levels and hepatic protein synthesis, the latter of which appeared to be mammalian target of rapamycin independent and was associated with increased hepatic tumor necrosis factor alpha messenger RNA content.

Conclusions: Without increasing mortality, the ZM state is associated with a more disordered acute systemic inflammatory response and persistence or enhancement of acute phase responses within the liver parenchyma.
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http://dx.doi.org/10.1016/j.jss.2016.10.020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5412598PMC
April 2017

ZnT2-Mediated Zinc Import Into Paneth Cell Granules Is Necessary for Coordinated Secretion and Paneth Cell Function in Mice.

Cell Mol Gastroenterol Hepatol 2016 May 8;2(3):369-383. Epub 2016 Jan 8.

Department of Surgery, Penn State Hershey College of Medicine, Hershey, Pennsylvania; Department of Cellular and Molecular Physiology, Penn State Hershey College of Medicine, Hershey, Pennsylvania; Department of Pharmacology, Penn State Hershey College of Medicine, Hershey, Pennsylvania.

Background & Aims: Defects in Paneth cell (PC) function are associated with microbial dysbiosis and intestinal inflammation. PC granules contain antimicrobial peptides, cytokines, and substantial stores of zinc (Zn). We hypothesized that Zn, transported into the granule through the Zn transporter (ZnT)2, is critical for signature PC functions.

Methods: ZnT2 was localized to PC granules using immunofluorescence and sucrose gradient fractionation in wild-type () mice, and consequences of ZnT2 loss were characterized in ZnT2 knockout () mice. Terminal ilea were harvested for immunofluorescence, electron microscopy, and fluorescent imaging with the Zn reporter Zinpyr-1. Alterations in fecal microbiota were characterized using 16s ribosomal RNA sequencing. PC degranulation, bacterial translocation, cytokine response to endotoxin lipopolysaccharide, crypt viability after exposure to the oxidant monochloramine (NHCl), and bactericidal activity of luminal contents of terminal ilea against enteropathogenic were assessed.

Results: ZnT2 was localized to the membrane of PC granules. In mice, spontaneous degranulation was observed more frequently than among mice. Secretory granules were hypodense with less active lysozyme, and there was evidence of autophagosome accumulation and granule degradation in PCs from mice. Gut microbiota of mice were enriched in S24-7 and relatively depleted of species commonly found in mice. Evidence of PC dysfunction in mice included impaired granule secretion and increased inflammatory response to lipopolysaccharide, less bactericidal activity, and greater susceptibility to cell death from NHCl.

Conclusions: ZnT2 is critical for Zn import into PC granules, and the inability to import Zn leads to profound defects in PC function and uncoordinated granule secretion.
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http://dx.doi.org/10.1016/j.jcmgh.2015.12.006DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5042355PMC
May 2016

Oral omega-3 fatty acids promote resolution in chemical peritonitis.

J Surg Res 2016 11 17;206(1):190-198. Epub 2016 Jun 17.

Department of Surgery, Hershey, Pennsylvania; Department of Cellular and Molecular Physiology, Hershey, Pennsylvania. Electronic address:

Background: Recent studies suggest that purified omega-3 fatty acids may attenuate acute inflammation and hasten the transition to healing. In this study, we tested the hypothesis that pretreatment with omega-3-rich fish oil (FO) would promote resolution of peritoneal inflammation through production of specific lipid mediators.

Methods: C57/BL6 mice were given a daily 200-μL oral gavage of saline (CTL) or FO (1.0-1.5 g/kg/d docosahexaenoic acid and 1.3-2.0 g/kg/d eicosapentaenoic acid) for 7 d before chemical peritonitis was induced with thioglycollate. Peritoneal lavage fluid was collected before induction and at days 2 and 4 after peritonitis onset. Prostaglandin E2 (PGE2), Leukotriene B4 (LTB4), Resolvin D1 (RvD1), and the composition of immune cell populations were examined in peritoneal lavage exudates. Cells harvested from the peritoneum were assessed for macrophage differentiation markers, phagocytosis, and lipopolysaccharide-induced cytokine secretion profiles (interleukin [IL]-6, IL-10, IL-1β, TNFα).

Results: The ratio of RvD1 to pro-inflammatory PGE2 and LTB4 was increased in the peritoneal cavity of FO-supplemented animals. FO induced a decrease in the number of monocytes in the lavage fluid, with no change in the number of macrophages, neutrophils, or lymphocytes. Macrophage phagocytosis and M1/M2 messenger RNA markers were unchanged by FO with the exception of decreased PPARγ expression. FO increased ex vivo TNFα secretion after stimulation with lipopolysaccharide.

Conclusions: Our findings provide evidence that nutraceutically relevant doses of FO supplements given before and during chemical peritonitis shift the balance of lipid mediators towards a proresolution, anti-inflammatory state without drastically altering the number or phenotype of local innate immune cell populations.
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http://dx.doi.org/10.1016/j.jss.2016.06.036DOI Listing
November 2016

Marginal dietary zinc deprivation augments sepsis-induced alterations in skeletal muscle TNF-α but not protein synthesis.

Physiol Rep 2016 11;4(21)

Department of Surgery, College of Medicine, Pennsylvania State University, Hershey, Pennsylvania

Severe zinc deficiency is associated with an increased systemic inflammatory response and mortality after sepsis. However, the impact of mild zinc deficiency, which is more common in populations with chronic illnesses and sepsis, is unknown. In this study, we hypothesized that marginal dietary Zn deprivation (ZM) would amplify tissue inflammation and exacerbate the sepsis-induced decrease in muscle protein synthesis. Adult male C57BL/6 mice were fed a zinc-adequate (ZA) or ZM diet (30 or 10 mg Zn/kg, respectively) over 4 weeks, peritonitis was induced by cecal ligation and puncture (CLP), and mice were examined at either 24 h (acute) or 5 days (chronic) post-CLP Acute sepsis decreased the in vivo rate of skeletal muscle protein synthesis and the phosphorylation of the mTOR substrate 4E-BP1. Acutely, sepsis increased TNF-α and IL-6 mRNA in muscle, and the increase in TNF-α was significantly greater in ZM mice. However, muscle protein synthesis and 4E-BP1 phosphorylation returned to baseline 5 days post-CLP in both ZA and ZM mice. Protein degradation via markers of the ubiquitin proteasome pathway was increased in acute sepsis, yet only MuRF1 mRNA was increased in chronic sepsis and ZM amplified this elevation. Our data suggest that mild zinc deficiency increases TNF-α in muscle acutely after sepsis but does not significantly modulate the rate of muscle protein synthesis.
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http://dx.doi.org/10.14814/phy2.13017DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5112495PMC
November 2016

Biological underpinnings of breastfeeding challenges: the role of genetics, diet, and environment on lactation physiology.

Am J Physiol Endocrinol Metab 2016 08 28;311(2):E405-22. Epub 2016 Jun 28.

Departments of Cellular and Molecular Physiology, Pharmacology, and Surgery, Pennsylvania State Hershey College of Medicine, Hershey, Pennsylvania; and Department of Nutritional Sciences, The Pennsylvania State University, University Park, Pennsylvania

Lactation is a dynamic process that has evolved to produce a complex biological fluid that provides nutritive and nonnutritive factors to the nursing offspring. It has long been assumed that once lactation is successfully initiated, the primary factor regulating milk production is infant demand. Thus, most interventions have focused on improving breastfeeding education and early lactation support. However, in addition to infant demand, increasing evidence from studies conducted in experimental animal models, production animals, and breastfeeding women suggests that a diverse array of maternal factors may also affect milk production and composition. In this review, we provide an overview of our current understanding of the role of maternal genetics and modifiable factors, such as diet and environmental exposures, on reproductive endocrinology, lactation physiology, and the ability to successfully produce milk. To identify factors that may affect lactation in women, we highlight some information gleaned from studies in experimental animal models and production animals. Finally, we highlight the gaps in current knowledge and provide commentary on future research opportunities aimed at improving lactation outcomes in breastfeeding women to improve the health of mothers and their infants.
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http://dx.doi.org/10.1152/ajpendo.00495.2015DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5005964PMC
August 2016

Erratum to: Exome Sequencing of SLC30A2 Identifies Novel Loss- and Gain-of-Function Variants Associated with Breast Cell Dysfunction.

J Mammary Gland Biol Neoplasia 2016 12;21(3-4):149

Department of Cellular and Molecular Physiology, The Pennsylvania State University Hershey College of Medicine, Hershey, PA, USA.

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http://dx.doi.org/10.1007/s10911-016-9357-4DOI Listing
December 2016

Molecular regulation of lactation: The complex and requisite roles for zinc.

Arch Biochem Biophys 2016 Dec 6;611:86-92. Epub 2016 Apr 6.

Departments of Cellular and Molecular Physiology, Penn State Hershey College of Medicine, Hershey, PA 17033, USA; Departments of Pharmacology, Penn State Hershey College of Medicine, Hershey, PA 17033, USA; Departments of Surgery, Penn State Hershey College of Medicine, Hershey, PA 17033, USA. Electronic address:

Lactation provides many health benefits to the nursing infant and breastfeeding mother. In order to successfully breastfeed, the mammary gland must expand and differentiate to activate numerous processes that regulate milk production and secretion. This involves a complex series of molecular, biochemical and cellular events driven largely by lactogenic hormones. Recent advances implicate zinc as a critical modulator of mammary gland function. Here, we provide an overview of our current understanding of the role and regulation of zinc in promoting proliferation, differentiation and secretion in the mammary gland during lactation, and highlight critical gaps in knowledge.
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http://dx.doi.org/10.1016/j.abb.2016.04.002DOI Listing
December 2016

Subtype-specific accumulation of intracellular zinc pools is associated with the malignant phenotype in breast cancer.

Mol Cancer 2016 Jan 5;15. Epub 2016 Jan 5.

The Interdisciplinary Graduate Program in Physiology, Penn State Hershey College of Medicine, Hershey, PA, 17033, USA.

Background: Zinc (Zn) hyper-accumulates in breast tumors and malignant cell lines compared to normal mammary epithelium. The mechanisms responsible for Zn accumulation and the consequence of Zn dysregulation are poorly understood.

Methods: Microarrays were performed to assess differences in the expression of Zn transporters and metallothioneins (MTs) in human breast tumors and breast cancer cell lines. Real-time PCR and immunoblotting were employed to profile Zn transporter expression in representative luminal (T47D), basal (MDA-MB-231), and non-malignant (MCF10A) cell lines. Zn distribution in human tumors was assessed by X-ray fluorescence imaging. Zn distribution and content in cell lines was measured using FluoZin-3 imaging, and quantification and atomic absorption spectroscopy. Functional consequences of ZnT2 over-expression in MDA-MB-231 cells including invasion, proliferation, and cell cycle were measured using Boyden chambers, MTT assays, and flow cytometry, respectively.

Results: Gene expression profiling of human breast tumors and breast cancer cell lines identified subtype-specific dysregulation in the Zn transporting network. X-ray fluorescence imaging of breast tumor tissues revealed Zn hyper-accumulation at the margins of Luminal breast tumors while Zn was more evenly distributed within Basal tumors. While both T47D and MDA-MB-231 cells hyper-accumulated Zn relative to MCF10A cells, T47D cells accumulated 2.5-fold more Zn compared to MDA-MB-231 cells. FluoZin-3 imaging indicated that Zn was sequestered into numerous large vesicles in T47D cells, but was retained in the cytoplasm and found in fewer and larger, amorphous sub-cellular compartments in MDA-MB-231 cells. The differences in Zn localization mirrored the relative abundance of the Zn transporter ZnT2; T47D cells over-expressed ZnT2, whereas MDA-MB-231 cells did not express ZnT2 protein due to proteasomal degradation. To determine the functional relevance of the lack of ZnT2 in MDA-MB-231cells, cells were transfected to express ZnT2. ZnT2 over-expression led to Zn vesicularization, shifts in cell cycle, enhanced apoptosis, and reduced proliferation and invasion.

Conclusions: This comprehensive analysis of the Zn transporting network in malignant breast tumors and cell lines illustrates that distinct subtype-specific dysregulation of Zn management may underlie phenotypic characteristics of breast cancers such as grade, invasiveness, metastatic potential, and response to therapy.
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http://dx.doi.org/10.1186/s12943-015-0486-yDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4700748PMC
January 2016

Marginal zinc intake reduces the protective effect of lactation on mammary gland carcinogenesis in a DMBA-induced tumor model in mice.

Oncol Rep 2016 Mar 22;35(3):1409-16. Epub 2015 Dec 22.

Department of Surgery, The Pennsylvania State University Hershey College of Medicine, Hershey, PA 17033, USA.

Breastfeeding can reduce breast cancer risk; however, unknown factors modify this protective effect. Zinc (Zn) modulates an array of cellular functions including oxidative stress, cell proliferation, motility and apoptosis. Marginal Zn intake is common in women and is associated with breast cancer. We reported that marginal Zn intake in mice leads to mammary gland hypoplasia and hallmarks of pre-neoplastic lesions. In the present study, we tested the hypothesis that marginal Zn intake confounds the protective effect of lactation on breast cancer. Nulliparous mice fed control (ZA, 30 mg Zn/kg) or a marginal Zn diet (ZD, 15 mg Zn/kg), were bred and offspring were weaned naturally. Post-involution, mice were gavaged with corn oil or 7,12-dimethylbenz(a)anthracene (DMBA, 1 mg/wk for 4 weeks) and tumor development was monitored. A ZD diet led to insufficient involution, increased fibrosis and oxidative stress. Following DMBA treatment, mice fed ZD had higher oxidative stress in mammary tissue that correlated with reduced levels of peroxiredoxin-1 and p53 and tended to have shorter tumor latency and greater incidence of non-palpable tumors. In summary, marginal Zn intake creates a toxic mammary gland microenvironment and abrogates the protective effect of lactation on carcinogenesis.
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http://dx.doi.org/10.3892/or.2015.4508DOI Listing
March 2016

Impaired recovery from peritoneal inflammation in a mouse model of mild dietary zinc restriction.

Mol Nutr Food Res 2016 Mar 2;60(3):672-81. Epub 2016 Feb 2.

Departments of Surgery, Hershey, PA, USA.

Scope: Mild dietary zinc (Zn) deficiency is wide-spread in human populations, but the effect on Zn-dependent processes of immune function and healing are not well understood. The consequences of mild dietary Zn restriction were examined in two mouse models of inflammation and recovery.

Methods And Results: Male C57BL/6 mice were fed a Zn adequate diet (ZA, 30 mg Zn/kg diet), or diets containing sub-optimal Zn levels (ZM, 15 mg Zn/kg diet; ZD, 10 mg Zn/kg diet) for 30 days before a thioglycollate peritonitis challenge. Plasma lipid profiles were distinct, with greater Zn restriction resulting in a greater impact on metabolites. The milder ZM diet was selected for immune studies. Peritoneal macrophages from ZM mice displayed increased phagocytosis and amplified pro-inflammatory cytokine (IL-1β, IL-6, and TNFα) release compared to ZA, at baseline and after a secondary LPS challenge. Splenocytes isolated from ZM mice displayed an increase in IL-6 and a reduction in anti-inflammatory IL-4 compared to ZA. Cytokine levels in plasma were unaltered. Following mechanical manipulation of the intestines to induce ileus, ZM mice had delayed intestinal transit compared to ZA.

Conclusion: Mild Zn deficiency enhances local inflammatory responses, amplifying macrophage functions and delaying recovery from acute insults within the peritoneum.
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http://dx.doi.org/10.1002/mnfr.201500688DOI Listing
March 2016

ZnT4 (SLC30A4)-null ("lethal milk") mice have defects in mammary gland secretion and hallmarks of precocious involution during lactation.

Am J Physiol Regul Integr Comp Physiol 2016 Jan 4;310(1):R33-40. Epub 2015 Nov 4.

Department of Cellular and Molecular Physiology, Penn State Hershey College of Medicine, Hershey, Pennsylvania; Department of Pharmacology, Penn State Hershey College of Medicine, Hershey, Pennsylvania; Department of Surgery, Penn State Hershey College of Medicine, Hershey, Pennsylvania; and Department of Nutritional Sciences, The Pennsylvania State University, University Park, Pennsylvania

During lactation, highly specialized secretory mammary epithelial cells (MECs) produce and secrete huge quantities of nutrients and nonnutritive factors into breast milk. The zinc (Zn) transporter ZnT4 (SLC30A4) transports Zn into the trans-Golgi apparatus for lactose synthesis, and across the apical cell membrane for efflux from MECs into milk. This is consistent with observations in "lethal milk" (lm/lm) mice, which have a truncation mutation in SLC30A4, and present with not only low milk Zn concentration, but also smaller mammary glands, decreased milk volume, and lactation failure by lactation day 2. However, the molecular underpinnings of these defects are not understood. Here, we used lactating C57BL/6J(lm/lm) (ZnT4-null) mice to explore the consequences of a ZnT4-null phenotype on mammary gland function during early lactation. Lactating C57BL/6J(lm/lm) mice had significantly fewer, smaller, and collapsed alveoli comprising swollen, lipid-filled MECs during early lactation. These defects were associated with decreased Akt expression and STAT5 activation, indicative of defects in MEC secretion. In addition, increased expression of ZnT2, TNF-α, and cleaved e-cadherin concomitant with increased activation of STAT3 implicated the loss of ZnT4 in precocious activation of involution. Collectively, our study indicates that the loss of ZnT4 has profound consequences on MEC secretion and may promote tissue remodeling in the mammary gland during early lactation.
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http://dx.doi.org/10.1152/ajpregu.00315.2014DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4796636PMC
January 2016

Exome Sequencing of SLC30A2 Identifies Novel Loss- and Gain-of-Function Variants Associated with Breast Cell Dysfunction.

J Mammary Gland Biol Neoplasia 2015 Dec 21;20(3-4):159-72. Epub 2015 Aug 21.

Department of Cellular and Molecular Physiology, The Pennsylvania State University Hershey College of Medicine, Hershey, PA, USA.

The zinc (Zn) transporter ZnT2 (SLC30A2) is expressed in specialized secretory cells including breast, pancreas and prostate, and imports Zn into mitochondria and vesicles. Mutations in SLC30A2 substantially reduce milk Zn concentration ([Zn]) and cause severe Zn deficiency in exclusively breastfed infants. Recent studies show that ZnT2-null mice have low milk [Zn], in addition to profound defects in mammary gland function during lactation. Here, we used breast milk [Zn] to identify novel non-synonymous ZnT2 variants in a population of lactating women. We also asked whether specific variants induce disturbances in intracellular Zn management or cause cellular dysfunction in mammary epithelial cells. Healthy, breastfeeding women were stratified into quartiles by milk [Zn] and exonic sequencing of SLC30A2 was performed. We found that 36% of women tested carried non-synonymous ZnT2 variants, all of whom had milk Zn levels that were distinctly above or below those in women without variants. We identified 12 novel heterozygous variants. Two variants (D(103)E and T(288)S) were identified with high frequency (9 and 16%, respectively) and expression of T(288)S was associated with a known hallmark of breast dysfunction (elevated milk sodium/potassium ratio). Select variants (A(28)D, K(66)N, Q(71)H, D(103)E, A(105)P, Q(137)H, T(288)S and T(312)K) were characterized in vitro. Compared with wild-type ZnT2, these variants were inappropriately localized, and most resulted in either 'loss-of-function' or 'gain-of-function', and altered sub-cellular Zn pools, Zn secretion, and cell cycle check-points. Our study indicates that SLC30A2 variants are common in this population, dysregulate Zn management and can lead to breast cell dysfunction. This suggests that genetic variation in ZnT2 could be an important modifier of infant growth/development and reproductive health/disease. Importantly, milk [Zn] level may serve as a bio-reporter of breast function during lactation.
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http://dx.doi.org/10.1007/s10911-015-9338-zDOI Listing
December 2015

Obesity-Induced Inflammation Is Associated with Alterations in Subcellular Zinc Pools and Premature Mammary Gland Involution in Lactating Mice.

J Nutr 2015 Sep 22;145(9):1999-2005. Epub 2015 Jul 22.

Department of Nutritional Sciences, The Pennsylvania State University, University Park, PA; and Departments of Cell and Molecular Physiology, Pharmacology, and Surgery, Penn State Hershey College of Medicine, Hershey, PA

Background: Lactation failure is common in overweight and obese women; however, the precise mechanism remains unknown.

Objective: We tested the hypothesis that obesity-induced inflammation in the mammary gland (MG) redistributes subcellular zinc pools to promote cell death of mammary epithelial cells (MECs) and premature involution.

Methods: Female DBA/2J mice were fed a high-fat (obese; 45% kcal from fat, n = 60) or control diet (lean; 10% kcal from fat, n = 50) for 5 wk and bred. MG cytokines and macrophage infiltration were determined by reverse transcriptase-polymerase chain reaction and F4/80 staining, respectively. Zinc concentration was analyzed by atomic absorption spectroscopy, and zinc transporters and markers of endoplasmic reticulum (ER) stress, autophagy, and involution were measured by immunoblot. To confirm effects of inflammation, tumor necrosis factor-α (TNF) or vehicle was injected into adjacent MGs of lean lactating C57BL/6 mice (n = 5) and cultured MECs (HC11 cells) were treated with TNF in vitro.

Results: Seventy-seven percent of obese mice failed to lactate (lean: 39%; P < 0.001). Obese mice capable of lactating had greater macrophage infiltration (obese: 135 ± 40.4 macrophages/mm(2); lean: 63.8 ± 8.9 macrophages/mm(2); P < 0.001) and elevated TNF expression (P < 0.05), concurrent with lower zrt- irt-like protein 7 abundance (P < 0.05) and higher ER zinc concentration (obese: 0.36 ± 0.004 μg Zn/mg protein; lean: 0.30 ± 0.02 μg Zn/mg protein; P < 0.05) compared with lean mice. Heat shock protein 5 (HSPA5) expression (P < 0.05) was suppressed in the MG of obese mice, which was consistent with HSPA5 suppression in TNF-injected MGs (P < 0.01) and MECs treated with TNF in vitro (P < 0.01). Moreover, obesity increased lysosomal activity (P < 0.05) and autophagy in the MG, which corresponded to increased zinc transporter 2 abundance and lysosomal zinc concentration compared with lean mice (obese: 0.20 ± 0.02 μg Zn/mg protein; lean: 0.14 ± 0.01 μg Zn/mg protein; P < 0.05). Importantly, MGs of obese mice exhibited markers of apoptosis (P = 0.05) and involution (P < 0.01), which were not observed in lean mice.

Conclusions: Diet-induced obesity created a proinflammatory MG microenvironment in mice, which was associated with zinc-mediated ER stress and autophagy and the activation of premature involution.
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http://dx.doi.org/10.3945/jn.115.214122DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4548167PMC
September 2015

Essential Role for Zinc Transporter 2 (ZnT2)-mediated Zinc Transport in Mammary Gland Development and Function during Lactation.

J Biol Chem 2015 May 7;290(21):13064-78. Epub 2015 Apr 7.

From the Interdisciplinary Graduate Physiology Program and Department of Nutritional Sciences, The Pennsylvania State University, University Park, Pennsylvania 16802, Departments of Cellular and Molecular Physiology, Surgery, Penn State Hershey College of Medicine, Hershey, Pennsylvania 17033, Pharmacology, and

The zinc transporter ZnT2 (SLC30A2) imports zinc into vesicles in secreting mammary epithelial cells (MECs) and is critical for zinc efflux into milk during lactation. Recent studies show that ZnT2 also imports zinc into mitochondria and is expressed in the non-lactating mammary gland and non-secreting MECs, highlighting the importance of ZnT2 in general mammary gland biology. In this study we used nulliparous and lactating ZnT2-null mice and characterized the consequences on mammary gland development, function during lactation, and milk composition. We found that ZnT2 was primarily expressed in MECs and to a limited extent in macrophages in the nulliparous mammary gland and loss of ZnT2 impaired mammary expansion during development. Secondly, we found that lactating ZnT2-null mice had substantial defects in mammary gland architecture and MEC function during secretion, including fewer, condensed and disorganized alveoli, impaired Stat5 activation, and unpolarized MECs. Loss of ZnT2 led to reduced milk volume and milk containing less protein, fat, and lactose compared with wild-type littermates, implicating ZnT2 in the regulation of mammary differentiation and optimal milk production during lactation. Together, these results demonstrate that ZnT2-mediated zinc transport is critical for mammary gland function, suggesting that defects in ZnT2 not only reduce milk zinc concentration but may compromise breast health and increase the risk for lactation insufficiency in lactating women.
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http://dx.doi.org/10.1074/jbc.M115.637439DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4505563PMC
May 2015

TNFα Post-Translationally Targets ZnT2 to Accumulate Zinc in Lysosomes.

J Cell Physiol 2015 Oct;230(10):2345-50

Department of Nutritional Sciences, The Pennsylvania State University, University Park, Pennsylvania.

Mammary epithelial cells undergo widespread lysosomal-mediated cell death (LCD) during early mammary gland involution. Recently, we demonstrated that tumor necrosis factor-α (TNFα), a cytokine released during early involution, redistributes the zinc (Zn) transporter ZnT2 to accumulate Zn in lysosomes and activate LCD and involution. The objective of this study is to determine how TNFα retargets ZnT2 to lysosomes. We tested the hypothesis that TNFα signaling dephosphorylates ZnT2 to uncover a highly conserved dileucine motif (L294L) in the C-terminus of ZnT2, allowing adaptor protein complex-3 (AP-3) to bind and traffic ZnT2 to lysosomes. Confocal micrographs showed that TNFα redistributed wild-type (WT) ZnT2 from late endosomes (Pearson's coefficient = 0.202 ± 0.05 and 0.097 ± 0.03; P<0.05) to lysosomes (0.292 ± 0.03 and 0.649 ± 0.03; P<0.0001), which increased lysosomal Zn (P<0.0001) and activated LCD (P<0.0001) compared to untreated cells. Mutation of the dileucine motif (L294V) eliminated the ability of TNFα to redistribute ZnT2 from late endosomes to lysosomes, increase lysosomal Zn, or activate LCD. Moreover, TNFα increased (P<0.05) AP-3 binding to wt ZnT2 but not to L294V immunoprecipitates. Finally, using phospho- and dephospho-mimetics of predicted phosphorylation sites (T281, T288, and S296), we found that dephosphorylated S296 was required to target ZnT2 to accumulate Zn in lysosomes and activate LCD. Our findings suggest that women with variation in the C-terminus of ZnT2 may be at risk for inadequate involution and breast disease due the inability to traffic ZnT2 to lysosomes.
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http://dx.doi.org/10.1002/jcp.24992DOI Listing
October 2015

ZnT2 is a critical mediator of lysosomal-mediated cell death during early mammary gland involution.

Sci Rep 2015 Jan 26;5:8033. Epub 2015 Jan 26.

1] Department of Nutritional Sciences, The Pennsylvania State University, University Park, PA, USA [2] Department of Surgery, Penn State Hershey College of Medicine, Hershey, PA, USA [3] Department of and Cell and Molecular Physiology, Penn State Hershey College of Medicine, Hershey, PA, USA [4] Department of Pharmacology, Penn State Hershey College of Medicine, Hershey, PA, USA.

Mammary gland involution is the most dramatic example of physiological cell death. It occurs through an initial phase of lysosomal-mediated cell death (LCD) followed by mitochondrial-mediated apoptosis. Zinc (Zn) activates both LCD and apoptosis in vitro. The Zn transporter ZnT2 imports Zn into vesicles and mitochondria and ZnT2-overexpression activates cell death in mammary epithelial cells (MECs). We tested the hypothesis that ZnT2-mediated Zn transport is critical for mammary gland involution in mice. Following weaning, ZnT2 abundance increased in lysosomes and mitochondria, which paralleled Zn accumulation in each of these organelles. Adenoviral expression of ZnT2 in lactating mouse mammary glands in vivo increased Zn in lysosomes and mitochondria and activated LCD and apoptosis, promoting a profound reduction in MECs and alveoli. Injection of TNFα, a potent activator of early involution, into the mammary gland fat pads of lactating mice increased ZnT2 and Zn in lysosomes and activated premature involution. Exposure of cultured MECs to TNFα redistributed ZnT2 to lysosomes and increased lysosomal Zn, which activated lysosomal swelling, cathepsin B release, and LCD. Our data implicate ZnT2 as a critical mediator of cell death during involution and importantly, that as an initial involution signal, TNFα redistributes ZnT2 to lysosomes to activate LCD.
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http://dx.doi.org/10.1038/srep08033DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4306139PMC
January 2015

Dietary Iron and Colorectal Cancer Risk: A Review of Human Population Studies.

Crit Rev Food Sci Nutr 2016 ;56(6):1012-20

e Department of Epidemiology , Rollins School of Public Health and Winship Cancer Institute, Emory University , Atlanta , Georgia , USA.

Iron is an essential micronutrient that is involved in many redox processes and serves as an integral component in various physiological functions. However, excess iron can cause tissue damage through its pro-oxidative effects, potentiating the development of many diseases such as cancer through the generation of reactive oxidative species. The two major forms of iron in the diet are heme and nonheme iron, both of which are found in several different foods. In addition to natural food sources, intake of nonheme iron may also come from fortified foods or in supplement form. This review summarizes the results of human population studies that have examined the role of dietary iron (heme and nonheme), heme iron alone, and iron from supplements in colorectal carcinogenesis.
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http://dx.doi.org/10.1080/10408398.2012.749208DOI Listing
January 2017

Paradoxical zinc toxicity and oxidative stress in the mammary gland during marginal dietary zinc deficiency.

Reprod Toxicol 2015 Jul 1;54:84-92. Epub 2014 Aug 1.

Department of Nutritional Sciences, The Pennsylvania State University, United States; Interdisciplinary Graduate Program in Physiology, The Pennsylvania State University, United States; Department of Surgery, Penn State Hershey College of Medicine, United States; Department of Cell and Molecular Physiology, Penn State Hershey College of Medicine, United States. Electronic address:

Zinc (Zn) regulates numerous cellular functions. Zn deficiency is common in females; ∼80% of women and 40% of adolescent girls consume inadequate Zn. Zn deficiency enhances oxidative stress, inflammation and DNA damage. Oxidative stress and inflammation is associated with breast disease. We hypothesized that Zn deficiency increases oxidative stress in the mammary gland, altering the microenvironment and architecture. Zn accumulated in the mammary glands of Zn deficient mice and this was associated with macrophage infiltration, enhanced oxidative stress and over-expression of estrogen receptor α. Ductal and stromal hypercellularity was associated with aberrant collagen deposition and disorganized e-cadherin. Importantly, these microenvironmental alterations were associated with substantial impairments in ductal expansion and mammary gland development. This is the first study to show that marginal Zn deficiency creates a toxic microenvironment in the mammary gland impairing breast development. These changes are consistent with hallmarks of potential increased risk for breast disease and cancer.
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http://dx.doi.org/10.1016/j.reprotox.2014.07.076DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4312747PMC
July 2015

Prolactin (PRL)-stimulated ubiquitination of ZnT2 mediates a transient increase in zinc secretion followed by ZnT2 degradation in mammary epithelial cells.

J Biol Chem 2014 Aug 11;289(34):23653-61. Epub 2014 Jul 11.

the Department of Nutritional Sciences, Pennsylvania State University, University Park, Pennsylvania 16802, and From the Departments of Cell and Molecular Physiology, Pharmacology, and Surgery, Penn State Hershey College of Medicine, Hershey, Pennsylvania 17033,

The zinc transporter ZnT2 imports zinc into secretory vesicles and regulates zinc export from the mammary epithelial cell. Mutations in ZnT2 substantially impair zinc secretion into milk. The lactogenic hormone prolactin (PRL) transcriptionally increases ZnT2 expression through the Jak2/STAT5 signaling pathway, increasing zinc accumulation in secretory vesicles and zinc secretion. Herein, we report that PRL post-translationally stimulated ZnT2 ubiquitination, which altered ZnT2 trafficking and augmented vesicular zinc accumulation and secretion from mammary epithelial cells in a transient manner. Ubiquitination then down-regulated zinc secretion by stimulating degradation of ZnT2. Mutagenesis of two N-terminal lysine residues (K4R and K6R) inhibited ZnT2 ubiquitination, vesicular zinc accumulation and secretion, and protein degradation. These findings establish that PRL post-translationally regulates ZnT2-mediated zinc secretion in a multifactorial manner, first by enhancing zinc accumulation in vesicles to transiently enhance zinc secretion and then by activating ubiquitin-dependent ZnT2 degradation. This provides insight into novel mechanisms through which ZnT2 and zinc transport is tightly regulated in mammary epithelial cells.
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http://dx.doi.org/10.1074/jbc.M113.531145DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4156067PMC
August 2014

Redistribution of tissue zinc pools during lactation and dyshomeostasis during marginal zinc deficiency in mice.

J Trace Elem Med Biol 2015 Jan 11;29:170-5. Epub 2014 Jun 11.

Department of Nutritional Sciences, The Pennsylvania State University, University Park, PA 16802, USA; Departments of Surgery and Cell and Molecular Physiology, Penn State Hershey College of Medicine, Hershey, PA 17033, USA. Electronic address:

Zinc (Zn) requirements are increased during lactation. Increased demand is partially met through increased Zn absorption from the diet. It is estimated that 60-80% of women of reproductive age are at risk for Zn deficiency due to low intake of bioavailable Zn and increased demands during pregnancy and lactation. How Zn is redistributed within the body to meet the demands of lactation, and how Zn deficiency affects this process, is not understood. Female C57bl/6J mice were fed a control (ZA; 30mg Zn/kg) or a marginally Zn deficient (ZD; 15mg Zn/kg) diet for 30 days prior to mating through mid-lactation and compared with nulliparous mice fed the same diets. While stomach and plasma Zn concentration increased during lactation in mice fed ZA, mice fed ZD had lower stomach Zn concentration and abrogated plasma Zn levels during lactation. Additionally, femur Zn decreased during lactation in mice fed ZA, while mice fed ZD did not experience this decrease. Furthermore, red blood cell, pancreas, muscle and mammary gland Zn concentration increased, and liver and adrenal gland Zn decreased during lactation, independent of diet, while kidney Zn concentration increased only in mice fed ZD. Finally, maternal Zn deficiency significantly increased the liver Zn concentration in offspring but decreased weight gain and survival. This study provides novel insight into how Zn is redistributed to meet the increased metabolic demands of lactation and how marginal Zn deficiency interferes with these homeostatic adjustments.
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http://dx.doi.org/10.1016/j.jtemb.2014.06.002DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4258524PMC
January 2015
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