In this study, the fabrication of biocompatible MXene-reinforced imprinted membranes (MX-AIM) capable of selectively capturing acetaminophen (APAP) molecules during filtration was demonstrated. The incorporation of MXene into a polymeric support membrane led to an overall improvement in both mechanical integrity and surface reactivity of the resulting composite, subsequently allowing efficient deposition of a molecularly imprinted polymer (MIP) layer on its surface. The synergy between the high surface reactivities of MXenes and the MIP layer rendered more adsorption sites, leading to the capture of approximately 32 % APAP on MX-AIM from a 200 ppm drug feed solution, compared to only -2 % for directly imprinted membranes. Additionally, these MXene-modified membranes exhibited high cell viabilities when tested against human monocytic cells, reaching up to >190 % cell proliferation. Overall, this research offers important insights on incorporating MXene nanofillers into membrane production, with the potential to overcome the challenges associated with applying the molecular imprinting technique to enhance the functionality of polymer materials.

In this study, three functionalities of superhydrophobicity, antibacterial activity and electromagnetic interference (EMI) shielding of cotton fabrics coated with Ag/PDMS were studied, and the role of coating composition was discussed. Special attention was paid to understanding the relationships between the surface roughness of coated fibres with the developed superhydrophobicity and antibacterial activity. The superhydrophobicity of fabrics was analysed based on water contact angle (WCA) and contact angle hysteresis (CAH) values while the antibacterial activity was tested against Escherichia coli ( E. coli ) and Staphylococcus aureus ( S. aureus ) bacteria. It was found that the surface roughness on fibres, which changed by altering the concentrations of Ag NPs (0.2–4 g/L) and PDMS (20 and 40 g/L), affected the obtained superhydrophobicity. The most superhydrophobic fabric (WCA 171°) was coated with a formulation containing Ag NPs (2 g/L), and PDMS (20 g/L), and had the highest surface roughness. Increasing the ingredients’ concentrations, however, deteriorated the optimum roughness as measured using a 3D Surface Laser Scanning Microscopy method. The coated fabrics showed efficacy against both types of bacteria, and it was confirmed that the Ag NPs content was the key factor in determining the antibacterial performance. Moreover, testing the EMI shielding performance of fabrics demonstrated that increasing the concentrations of Ag NPs and PDMS both enhanced the reflection coefficient (R) of fabrics against incident X-band electromagnetic (EM) wave by around >500%, which was due to the deposition of a higher amount of Ag NPs on fabrics. The superhydrophobicity of fabrics was durable after 1000 abrasion cycles, and the fabrics retained their antibacterial activity even after numerous washings. Graphical Abstract

Hemodialysis acts as an artificial kidney that selectively removes specific toxins, bio-compounds, or fluid from the main blood stream in a patient with kidney failure. The current process uses ultrafiltration-based membrane technology, where a semi-permeable material selectively extracts chemicals, such as uremic retention products, or remove excess water from blood by retaining certain compounds based on their size. As sugars, fats, proteins, biomolecules, cells, and platelets move into and across the tubular membrane in the hemodialysis process, the surface of the membrane begins to foul, which leads to major operational challenges that include sharp pressure drops with increasing operation times. The design of membranes with enhanced biocompatibility and anti-fouling properties is one avenue to increase the lifespan of the membrane used while facilitating the device operation and limiting the stress and discomfort of patients. This review presents interfacial interactions between blood components and membrane materials used in hemodialysis. The discussion analyzes the impacts of the hemodialyzer module design, membrane material morphology and surface chemistry on the long-term operation and performance of the hemodialyzers. Avenues for the development of next-generation-membrane-materials as well as new strategies to enhance the selective removal of toxic compounds from blood are also discussed.

Cotton fabrics with superhydrophobic, antibacterial, UV protection, and photothermal properties were developed using Ag/PDMS coatings, and the role of coating formulations on the obtained functionalities was studied. Specific attention was paid to understanding the relationships between the fabrics' superhydrophobicity and antibacterial activity against Escherichia coli (E. coli) bacteria. UV protection performance of Ag/PDMS coatings was thoroughly evaluated based on the variation of UV transmission rate through coated fabrics and photoinduced chemiluminescence spectra. Moreover, the effect of silver nanoparticles (Ag NPs) and PDMS on developing a photothermal effect on fabrics was discussed. It was found that the content of Ag NPs and PDMS played critical roles in determining the water contact angle (WCA) on modified fabrics. The largest WCA was 171.31°, which was durable even after numerous accelerated wash cycles and abrasions. Antibacterial activity of fabrics showed the positive effect of pure PDMS in bacterial growth inhibition. Moreover, it was found that the antibacterial performance was greatly affected by the content of Ag NPs loaded on fabrics rather than their superhydrophobic status. Moreover, increasing the content of Ag NPs boosted the UV protection level of fabrics, improved fabrics photostability, and reduced the UV transmission rate through fabrics. Testing the photothermal effect confirmed that the content of Ag NPs and PDMS both played prominent roles, where Ag acted as a photothermal agent and PDMS determined the NIR reflection rate from the coated surface. The modified fabrics were characterized using TGA, SEM, FTIR, and XRD techniques, and it was confirmed that using a higher amount of PDMS increased the amount of Ag NPs deposition on fabrics.

Lactation is a defining feature of mammals. The production of milk by the mammary gland during lactation is generally regarded as a mechanism required to supply nutrition to the neonate before it is able to digest other types of food. Increasing evidence suggests that components of milk have additional functions in addition to nutrition. Monotremes, marsupials and eutherians are all members of the class mammalia, but monotremes and marsupials have evolved a lactation pattern that is very different from eutherians. In eutherians, the gestation period is long relative to its lactation period. However, in marsupials and monotremes the gestation period is relatively short. As a result, the newborn is relatively small and undeveloped and most of the early development occurs postnatally. Eutherians produce milk of a constant composition after the expression of the initial colostrum, whereas marsupials such as the tammar wallaby (Macropus eugenii), have a short gestation, and give birth to a highly immature young and then commence a long lactation during which the milk production and composition progressively change in composition to suit the needs of the developing young. The comparative study of monotreme, marsupial and eutherian milk is of significant interest and may reveal bioactives required for developmental processes. These factors may have been either lost, down-regulated or altered/modified in eutherians since most development of eutherian young occurs in utero with nutrition and developmental signaling provided by the amniotic fluid and supplemented from the mother via the placenta.

This study presents a new approach to imparting the combined functionalities of superhydrophobicity, UV protection and personal thermal management via the photothermal effect to cotton fabrics. Surface modification of fabrics was carried out by applying coatings containing natural melanin (NM) particles extracted from yak hair and fluorine-free polydimethylsiloxane (PDMS) polymer via a dip-pad-dry-cure process. Different concentrations of NM particles were used in the coatings and the roles of each ingredient in achieving the desired functionalities were investigated. In addition, the relationship between NM content and the resultant durability of the coatings was explored based on the variations in superhydrophobicity, UV protection, photothermal performance, and color strength of the fabrics. The results demonstrated that the non-fluorinated NM/PDMS coatings were highly effective in developing a superhydrophobic cotton fabric with a water contact angle of 164?degrees. The NM-coated fabrics provided a rapid heating effect under near-infrared light and their temperature increased to 38.4-45.3 degrees C, which was up to 14.7 degrees C higher than that of pristine cotton. Furthermore, the application of NM particles resulted in an excellent UV protection performance and generated the UV protection factor level of 198.48 on cotton fabrics. Finally, the coated fabrics were highly breathable and showed high stability against 1000 abrasion cycles, 5 accelerated washing tests, and 70 h UV irradiation. These findings provide new pathways in the development of future functional textiles using user-friendly and biocompatible ingredients.

Assessing the role of lactogenic hormones in human mammary gland development is limited due to issues accessing tissue samples and so development of a human in vitro three-dimensional mammosphere model with functions similar to secretory alveoli in the mammary gland can aid to overcome this shortfall. In this study, a mammosphere model has been characterised using human mammary epithelial cells grown on either mouse extracellular matrix or agarose and showed insulin is essential for formation of mammospheres. Insulin was shown to up-regulate extracellular matrix genes. Microarray analysis of these mammospheres revealed an up-regulation of differentiation, cell-cell junctions, and cytoskeleton organisation functions, suggesting mammosphere formation may be regulated through ILK signalling. Comparison of insulin and IGF-1 effects on mammosphere signalling showed that although IGF-1 could induce spherical structures, the cells did not polarise correctly as shown by the absence of up-regulation of polarisation genes and did not induce the expression of milk protein genes. This study demonstrated a major role for insulin in mammary acinar development for secretory differentiation and function indicating the potential for reduced lactational efficiency in women with obesity and gestational diabetes.

A series of Zn-Al-Li alloys with potential application in bioresorbable implants were cast, thermomechanically processed and tested. The formation of secondary phases, such as LiZn4, LiZn3Al and Al3Li, contributed to both dynamic recrystallization and grain refinement of the matrix (η-phase) during the hot-extrusion process, leading to grain sizes as small as 1.75 μm for Zn-4Al-0.6Li alloy (wt%). This alloy exhibited an ultimate tensile strength (UTS) of 451 MPa, a total elongation of 46% and a corrosion rate of 60 μm/year in simulated body fluid. The grain refinement played a major role in increasing the strength, but it also weakened the basal texture and promoted non-basal slip and grain boundary sliding, thus contributing to the increased plastic deformation of the alloy. The corrosion rate was affected by a layer of zinc oxide and phosphate formed in the early stages of the immersion tests. The corrosion products protected the substrate and tended to reduce the corrosion rate over time. The developed Zn-4Al-0.6Li and Zn-6Al-0.4Li alloys which showed promising mechanical and corrosion properties appeared to be cytocompatible in the mouse fibroblast cell line and human umbilical mesenchymal stem cells making them promising candidates for bioresorbable stent and implant applications.

A series of quaternary Zn-Al-Cu-Li alloys with different weight fractions of Cu, Al, and Li were developed and investigated for potential application in high load bearing bioresorbable implants. The developed alloys provided various fractions of binary and ternary intermetallic structures, which resulted in formation of multiphase microstructures containing a zinc-rich η-phase and LiZn4 and CuZn4 phases. The intermetallic phases promoted grain refinement and a good combination of mechanical properties. The developed Zn-2Al-4Cu-0.6Li alloy showed strength and ductility close to commercially pure Ti alloys with a UTS value of ∼535 MPa and elongation of 37%. The examination of in vitro corrosion behavior of the developed alloys in the modified Hanks' solution revealed suitable corrosion rates (∼38.5 μm/year). The moderate corrosion rate was controlled by the formation of a homogeneous layer of stable corrosion products that protected the alloys from the corrosive environment, particularly in the late stages of immersion. The developed alloys with the most promising mechanical and corrosion properties appeared to be biocompatible to mouse fibroblast cells and human umbilical mesenchymal stem cells, making them suitable candidates for implant applications.

The rapid advances in technology for both evaluating and understanding the structure of animal genomes and their functional significance have presented opportunities for scientists to more clearly understand the complexity of bovine milk proteins and the control of their expression. Used in conjunction with proteomic databases, we can start to expand our knowledge of how milk proteins are processed into peptides, which represent much of the biological activity residing within colostrum and milk. The challenge then remains to translate this information into products that form the basis of a functional foods industry, helping to underpin the commercial viability of the dairy industry. In this chapter, we present a review of the current status of bovine milk genomics and functional genomics, and describe the roles, characteristics and key bioactivities of the major bovine milk proteins and their encrypted peptides. The application of these analytical tools to the full spectrum of lactation strategies adopted by eutherians, marsupials and monotremes to improve our understanding of the milk proteome is discussed.

Milk has many functions, ranging from the provision of factors crucial to the operation of the mammary gland and the development of the suckled young through to their protection from infection. Lactation evolved about 200 million years ago with the aplacental, egg-laying monotremes, but since that time there has been extensive adaptation to reproduction, including a large repertoire of lactation strategies. This chapter discusses three animal models with extreme adaptation to lactation, and examines the option of exploiting their comparative biology to identify milk protein bioactives that may have potential in functional foods or pharmaceuticals. The echidna (Tachyglossus aculeatus, a monotreme) has a fascinating combination of reptilian and mammalian characteristics. It retains a primitive component of reptilian reproduction in the form of laying shelled eggs, but it also has a prototherian lactation process. The tammar wallaby (Macropus eugenii, a marsupial) has adopted a reproductive strategy that includes a short gestation (26.5 days), birth of an immature young, and a relatively long lactation (300 days). The composition of milk changes progressively during the lactation cycle, and these changes in milk composition subsequently control development of the young. The tammar can also practice concurrent asynchronous lactation; the mother provides a concentrated milk for an older animal that is out of the pouch, and a dilute milk from an adjacent mammary gland for a newborn pouch young. The third study species, the Cape fur seal (Arctocephalus pusilluspusillus, a eutherian), has a lactation characterized by a repeated cycle of long at-sea foraging trips (up to 23 days) alternating with short suckling periods of 2–3 days ashore. Lactation almost ceases while the seal is offshore, but the mammary gland does not progress to involution and apoptosis.Technology platforms using genomics, proteomics, and bioinformatics have been used to exploit these models to identify milk bioactives. In addition, the availability of sequenced marsupial, dog, platypus, and bovine genomes permits rapid transfer of information to the cow to provide outcomes for the dairy industry.

Natural melanin is recognized as a biocompatible photothermal agent because of its biologically derived nature and efficient photothermal conversion ability. Here, yak hair melanin (YM) is added to polyurethane (PU) for the fabrication of NIR-photoresponsive shape memory implants. The in vitro toxicity of the YM/PU composites is carried out by exposing them to human mesenchymal stem cells (hMSCs) and mouse fibroblast (L929) cells lines for 24 h, while the in vivo toxicity is investigated by implanting the YM/PU composites in the mouse for two months. No significant differences on cell viability, blood chemistry, hematology, and histological results are observed between YM/PU composites and control groups, suggesting their excellent biocompatibility. The biostability of the YM/PU composites is confirmed by monitoring their in vitro degradation for 12 weeks. The YM/PU column implanted in the back subcutis or vagina of the mouse rapidly recovered to its original state within 60 s under a very low NIR laser (808 nm, 0.5 W/cm2) intensity, which is much lower than the general laser intensity for photothermal cancer therapy (1-2 W/cm2). This work confirms the applicability of the YM/PU composites as long-term implant materials and expedites the use of YM/PU composites as cost-effective candidates for biomedical applications.

Significantly preterm and low-birthweight (LBW) babies have diminished lung and gut development, generally fail to thrive, have increased mortality and higher frequency of mature-onset disease. Mothers often cannot breastfeed, and babies receive either formula or pasteurized donor milk, which may further limit the baby's recovery. New approaches are required to manage the early stages of neonatal development. The tammar wallaby, an Australian marsupial, has a short gestation and a simple placenta, and gives birth to an altricial young equivalent to a final trimester human embryo. The neonate remains in the pouch and attached to the teat for 100 days postpartum. The mother slows growth of the young and progressively changes the composition of the milk to deliver signals for organ development, including the lung and gut. This closely resembles the relationship between the human fetus and delivery of placental and uterine bioactives. Datasets comprised of differentially expressed genes coding for secreted proteins in early lactation in the tammar mammary gland have been compared to databases produced from human placenta, amniotic fluid, colostrum and milk to identify human homologues for the putative signaling molecules for organ development. These data will be used to develop milk fortifiers for treatment of preterm and LBW babies in both the developed and the developing world.

EchAMP, the tenth most abundant transcript expressed in the mammary gland of echidna, has in vitro broad-spectrum antibacterial effects. However, the effects of EchAMP on mastitis, a condition where inflammation is triggered following mammary gland infection, has not been investigated. To investigate the impact of EchAMP against mastitis, EchAMP transgenic mice were generated. In antibacterial assays, the whey fractions of milk from transgenic mice significantly reduced growth of Staphylococcus aureus, Bacillus subtilis, Escherichia coli and Pseudomonas aeruginosa compared with whey fractions from wildtype mice. Furthermore, a mastitis model created by infecting mammary gland with these four bacterial strains displayed a significant reduction in bacterial load in transgenic mice injected with S. aureus and B. subtilis. On further confirmation, histomorphologic analysis showed absence of necrosis and cell infiltration in the mammary glands of transgenic mice. To understand the role of EchAMP against inflammation, we employed an LPS-injected mastitis mouse model. LPS is known to induce phopshorylation of NF-κB and MAPK pathways, which in turn activate downstream proinflammatory signaling mediators, to promote inflammation. In LPS-treated EchAMP transgenic mice, phosphorylation levels of NF-κB, p38 and ERK1/2 were significantly downregulated. Furthermore, in mammary gland of transgenic mice, there was a significant downregulation of mRNA levels of proinflammatory cytokines, namely TNF-α, IL-6 and IL-1β. Taken together, these data suggest that EchAMP has an antiinflammatory response and is effective against S. aureus and B. subtilis. We suggest that EchAMP may be a potential prophylactic protein against mastitis in dairy animals by expressing this gene in their mammary gland.

BACKGROUND:Antibiotic resistance is a problem that necessitates the identification of new antimicrobial molecules. Milk is known to have molecules with antimicrobial properties (AMPs). Echidna Antimicrobial Protein (EchAMP) is one such lactation specific AMP exclusively found in the milk of Echidna, an egg-laying mammal geographically restricted to Australia and New Guinea. Previous studies established that EchAMP exhibits substantial bacteriostatic activity against multiple bacterial genera. However, the subsequent structural and functional studies were hindered due to the unavailability of pure protein.RESULTS:In this study, we expressed EchAMP protein using a heterologous expression system and successfully purified it to >95% homogeneity. The purified recombinant protein exhibits bacteriolytic activity against both Gram-positive and Gram-negative bacteria as confirmed by live-dead staining and scanning electron microscopy. Structurally, this AMP belongs to the family of intrinsically disordered proteins (IDPs) as deciphered by the circular-dichroism, tryptophan fluorescence, and NMR spectroscopy. Nonetheless, EchAMP has the propensity to acquire structure with amphipathic molecules, or membrane mimics like SDS, lipopolysaccharides, and liposomes as again observed through multiple spectroscopic techniques.CONCLUSIONS:Recombinant EchAMP exhibits broad-spectrum bacteriolytic activity by compromising the bacterial cell membrane integrity. Hence, we propose that this intrinsically disordered antimicrobial protein interact with the bacterial cell membrane and undergoes conformational changes to form channels in the membrane resulting in cell lysis.GENERAL SIGNIFICANCE:EchAMP, the evolutionarily conserved, lactation specific AMP from an oviparous mammal may find application as a broad-spectrum antimicrobial against pathogens that affect mammary gland or otherwise cause routine infections in humans and livestock.

BACKGROUND:After a short gestation, marsupials give birth to immature neonates with lungs that are not fully developed and in early life the neonate partially relies on gas exchange through the skin. Therefore, significant lung development occurs after birth in marsupials in contrast to eutherian mammals such as humans and mice where lung development occurs predominantly in the embryo. To explore the mechanisms of marsupial lung development in comparison to eutherians, morphological and gene expression analysis were conducted in the gray short-tailed opossum (Monodelphis domestica).RESULTS:Postnatal lung development of Monodelphis involves three key stages of development: (i) transition from late canalicular to early saccular stages, (ii) saccular and (iii) alveolar stages, similar to developmental stages overlapping the embryonic and perinatal period in eutherians. Differentially expressed genes were identified and correlated with developmental stages. Functional categories included growth factors, extracellular matrix protein (ECMs), transcriptional factors and signalling pathways related to branching morphogenesis, alveologenesis and vascularisation. Comparison with published data on mice highlighted the conserved importance of extracellular matrix remodelling and signalling pathways such as Wnt, Notch, IGF, TGFβ, retinoic acid and angiopoietin. The comparison also revealed changes in the mammalian gene expression program associated with the initiation of alveologenesis and birth, pointing to subtle differences between the non-functional embryonic lung of the eutherian mouse and the partially functional developing lung of the marsupial Monodelphis neonates. The data also highlighted a subset of contractile proteins specifically expressed in Monodelphis during and after alveologenesis.CONCLUSION:The results provide insights into marsupial lung development and support the potential of the marsupial model of postnatal development towards better understanding of the evolution of the mammalian bronchioalveolar lung.

Umbilical cord-blood derived hematopoietic stem cells (HSCs) are an attractive source for HSC transplantation as a means to treat hematological diseases. However, the small number of HSCs extracted from umbilical cord-blood contributes to poor clinical outcomes. Expansion techniques ex vivo designed to increase the number of cord-blood HSCs depend on replicating the stem cell niche to promote progenitor cell expansion without differentiation. Niche growth factors such as some members of the Angiopoietin-like protein family (ANGPTL-2, 3 and 5) have been shown to aid HSC ex vivo expansion. However, these are large glycosylated proteins that readily degrade and form aggregates, making them difficult to purify for use in ex vivo expansion protocols. Therefore, this study aimed to identify novel cytokines that mimic the expansion effects of ANGPTL-2, 3 and 5. Comparative protein sequence analysis of ANGPTL-2, 3 and 5 identified a single fibrinogen-like domain (FLD) common between these proteins, which was subsequently used to search the NCBI database for FLD-containing proteins. A number of proteins were identified and Ficolin-1 (FCN1) was selected based on protein similarity and FLD conservation. The effect of FCN1 on ex vivo expansion of CD34+ HSCs was tested and flow cytometry demonstrated an increase in the proportion of CD34+ CD45+ and CD34+ CD133+ HSCs after 16 days compared to expansion in the absence of FCN1. A purified peptide of the FLD region of FCN1 (201aa- 300aa) was also an effective expander, suggesting this region alone is likely responsible for the effects of FCN1 on HSC expansion. FCN1 belongs to a group of lectins that have a well-established role in the Complement signalling pathway as part of the innate immune defence mechanism. However, the HSC expansion mechanism of FCN1 remains unknown. RNA sequencing and pathway analysis were used to investigate the signalling pathway involved in FCN1-mediated HSC expansion across different time points. Other FLD-containing proteins (Fibroleukin, Intelectin and Tenascin XB) were also investigated. Expansion experiments showed that while Fibroleukin and Intelectin expanded CD34+ CD45+ and CD34+ CD133+ HSCs after 16 days, Tenascin XB did not. This suggested that subtle differences in FLD sequence or surrounding sequence may lead to differences in activity. This study demonstrated the significance of utilising cytokine that mimicked the stem cell niche for the purpose of HSC ex vivo expansion.

Objective Beta-casein is a major protein in breast milk and an important source for several bioactive peptides that are encrypted within the sequence. Beta-casomorphins (BCMs) are short-chain proteolytic peptides that are derived from the beta-casein protein and have opioid effects in newborns. Human milk is known to contain naturally occurring milk-protein-derived bioactive peptides but the identification of naturally occurring beta-casein-derived BCMs in human breast milk has been limited due to difficulties in the detection of BCM peptides, which are small and circulate in low concentrations. Methods The present study aimed to identify the naturally occurring BCM peptides from beta-casein in human breast milk using liquid chromatography-tandem mass spectrometry. The BCM peptides identified in the breast milk were analysed to predict the milk proteases responsible for the cleavage patterns using a computational tool EnzymePredictor. Results In-depth peptidomics analysis of breast milk samples that were collected at different lactation stages during human lactation revealed the presence of BCMs including BCM-8, -9, -10, and -11 as well as precursors and truncated forms of the original peptide, which suggests that milk protease activity in the mammary gland generates biologically relevant BCMs. Conclusions To our knowledge, this is the first report to describe the presence of naturally occurring human BCM-10 and -11 in breast milk. Our study provides evidence of beta-casein-derived BCM peptides in human milk before infant digestion. Proteases that are present in milk are likely specific in their proteolysis of beta-casein. The identified bioactive BCM-8, -9, -10, and -11 as well as the precursor peptides meet the structural requirements to elicit opioid, immunomodulatory, antioxidative, and satiety functions in newborns.

Extracellular matrix (ECM) plays an important role in the normal physiology of tissues and progression to disease.Earlier studies and our external microarray data analysis indicated that mammary matrix from involuting tissue showedupregulation of genes involved in ECM remodeling. The present study examines the fate of mammary and oral cancercells grown in the ECM from lactating mammary gland. Our findings show that non-tumorigenic cells, MCF10A andDOK cells did not proliferate but the tumorigenic and metastatic cells, SCC25 and MDA-MB-231, underwent apoptosiswhen grown on mammary ECM isolated from lactating mice. In addition, the cytokinesis marker, CEP55, was repressedin the oral and breast cancer cells. In contrast, these cells proliferated normally on mammary ECM isolated from miceundergoing involution. External microarray data analysis of mammary tissue further revealed over expression (~16 fold)of QSOX1 gene, which promotes cellular quiescence, in lactating mammary gland. A recent study has indicated thatQSOX1 overexpression in breast cancer cells led to reduced proliferation and tumorigenic properties. This extracellularprotein in mammary ECM may be responsible for reduced cellular proliferation. The present study has shown that ECMfrom lactating mammary gland can regulate signals to oral and breast cancer cells to halt cell division. This preliminaryobservation provided insights into the potential role of ECM factors present in lactating mammary gland as therapeutictargets to control cancer cell division. This preliminary study is an attempt to understand not only the requirement ofECM remodeling factors essential for the growth and survival of cancer cells but also the factors present in the lactationmatrix that simultaneously halts cell division and selectively inhibits the growth of cancer cells.

Monotreme lactation protein (MLP) is a recently identified protein with antimicrobial activity. It is present in the milk of monotremes and is unique to this lineage. To characterize MLP and to gain insight into the potential role of this protein in the evolution of lactation, the crystal structure of duck-billed platypus (Ornithorhynchus anatinus) MLP was determined at 1.82 Å resolution. This is the first structure to be reported for this novel, mammalian antibacterial protein. MLP was expressed as a FLAG epitope-tagged protein in mammalian cells and crystallized readily, with at least three space groups being observed (P1, C2 and P21). A 1.82 Å resolution native data set was collected from a crystal in space group P1, with unit-cell parameters a = 51.2, b = 59.7, c = 63.1 Å, α = 80.15, β = 82.98, γ = 89.27°. The structure was solved by SAD phasing using a protein crystal derivatized with mercury in space group C2, with unit-cell parameters a = 92.7, b = 73.2, c = 56.5 Å, β = 90.28°. MLP comprises a monomer of 12 helices and two short β-strands, with much of the N-terminus composed of loop regions. The crystal structure of MLP reveals no three-dimensional similarity to any known structures and reveals a heretofore unseen fold, supporting the idea that monotremes may be a rich source for the identification of novel proteins. It is hypothesized that MLP in monotreme milk has evolved to specifically support the unusual lactation strategy of this lineage and may have played a central role in the evolution of these mammals.