Gestational giant panda plasma metabolomics: amino acid metabolism characteristics may predict panda pregnancy outcomes.

In brief: The metabolic processes of the gestation period in pandas remain poorly understood. Our study comprehensively characterizes the metabolism of giant pandas during gestation and proposes arginine and histidine as potential novel biomarkers for detecting the pregnancy state of giant pandas. Abstract: There has been remarkable progress in the conservation and reproduction of giant pandas. However, the physiology of the gestation period in pandas remains poorly understood. The metabolic processes from estrus to pregnancy are dynamic and precisely regulated, playing a crucial role in pregnancy and related dysfunctions. In this study, we conducted a metabolomic analysis of 37 blood samples collected from pandas in estrus, acyclic, and potential pregnant states, employing rigorous screening to minimize the influence of diet. Our findings suggest that a reduced appetite can serve as an indicator for evaluating implantation time, representing a characteristic response to pregnancy and aiding in the prediction of delivery time in pregnant pandas. Metabolomic results indicate great metabolism variation from estrus to pregnancy, highlighting the association between amino acid metabolism and pregnancy outcomes. Compared to other pandas, individuals who successfully bred exhibit significantly elevated levels of arginine and histidine, even 2 months before experiencing a reduced appetite. Furthermore, the lipid profile undergoes distinct dynamic changes only in estrus samples. In summary, our study comprehensively characterizes the metabolism of giant pandas during gestation and proposes arginine and histidine as potential novel biomarkers for detecting the pregnancy state of giant pandas.

Blood transcriptome analysis revealed the immune changes and immunological adaptation of wildness training giant pandas.

The giant panda (Ailuropoda melanoleuca) is a global flagship species for biodiversity conservation. As the time for captive giant pandas to be released into the wild matures, wildness training is provided to allow adaptation to their natural environment. It is assumed that changes in the immune system would be integral in this adaptation from captive to wild, where many more pathogens would be encountered in their natural habitats. Therefore, this study aims to determine the expression changes of immune-related genes and their potential as immunoassay markers for adaptation monitoring in wildness training giant pandas, and then to understand the adaptation strategy of wildness training giant pandas to the wild environment, thereby improving the success rate of panda reintroduction. We obtained 300 differentially expressed genes (DEGs) by RNA-seq, with 239 up-regulated and 61 down-regulated DEGs in wildness training giant pandas compared to captive pandas. Functional enrichment analysis indicated that up-regulated DEGs were enriched in several immune-related terms and pathways. There were 21 immune-related DEGs, in which most of them were up-regulated in wildness training giant pandas, including several critical innate and cellular immune genes. IL1R2 was the most significantly up-regulated gene and is a signature of homeostasis within the immune system. In the protein-protein interaction (PPI) analysis, CXCL8, CXCL10, and CCL5 were identified as the hub immune genes. Our results suggested that wildness training giant pandas have stronger innate and cellular immunity than captive giant pandas, and we proposed that a gene set of CXCL8, CXCL10, CCL5, CD3D, NFKBIA, TBX21, IL12RB2, and IL1R2 may serve as potential immunoassay markers to monitor and assess the immune status of wildness training giant pandas. Our study offers the first insight into immune alterations of wildness training giant pandas, paving the way for monitoring and evaluating the immune status of giant pandas when reintroducing them into the wild.

ANALYSIS OF SERUM PROTEINS IN HEALTHY GIANT PANDAS (AILUROPODA MELANOLEUCA) UNDER MANAGED CARE.

Electrophoresis is a practical diagnostic tool for the identification of changes in serum protein fractions, which can be associated with a variety of diseases. Protein electrophoresis studies in Ursidae are limited, and currently no published fraction values are available for the giant panda (Ailuropoda melanoleuca). The aim of this study was to describe the serum protein fractions in the giant panda using both capillary zone electrophoresis (CZE) and standard agarose gel electrophoresis (AGE) techniques. Serum samples from nine healthy giant pandas (n = 19) were used for this study. Samples were evaluated using CZE and standard AGE. The CZE procedure successfully resolved serum proteins into seven fractions: prealbumin; albumin; and α1-, α2-, ß1-, ß2-, and γ-globulin; while AGE separated serum into only six protein fractions: prealbumin; albumin; α1-, α2-, and ß-globulins; and γ-globulin. These data will serve as a preliminary baseline for further studies and provide insight for the medical management of giant pandas.

A Translational Model for Venous Thromboembolism: MicroRNA Expression in Hibernating Black Bears.

BACKGROUND: Hibernating American black bears have significantly different clotting parameters than their summer active counterparts, affording them protection against venous thromboembolism during prolonged periods of immobility. We sought to evaluate if significant differences exist between the expression of microRNAs in the plasma of hibernating black bears compared with their summer active counterparts, potentially contributing to differences in hemostasis during hibernation. MATERIALS AND METHODS: MicroRNA sequencing was assessed in plasma from 21 American black bears in summer active (n = 11) and hibernating states (n = 10), and microRNA signatures during hibernating and active state were established using both bear and human genome. MicroRNA targets were predicted using messenger RNA (mRNA) transcripts from black bear kidney cells. In vitro studies were performed to confirm the relationship between identified microRNAs and mRNA expression, using artificial microRNA and human liver cells. RESULTS: Using the bear genome, we identified 15 microRNAs differentially expressed in the plasma of hibernating black bears. Of these microRNAs, three were significantly downregulated (miR-141-3p, miR-200a-3p, and miR-200c-3p), were predicted to target SERPINC1, the gene for antithrombin, and demonstrated regulatory control of the gene mRNA expression in cell studies. CONCLUSIONS: Our findings suggest that the hibernating black bears' ability to maintain hemostasis and achieve protection from venous thromboembolism during prolonged periods of immobility may be due to changes in microRNA signatures and possible upregulation of antithrombin expression.

The Fecal and Serum Metabolomics of Giant Pandas Based on Untargeted Metabolomics.

Little is comprehensively known or understood about giant panda fecal and serum metabolites, which could serve as important indicators of the physiological metabolism of giant pandas. Therefore, we determined the contents of fecal and serum metabolites of giant pandas based on an untargeted metabolome. Four hundred and 955 metabolites were detected in the feces and serum of giant panda, respectively. Glycerophospholipid and choline metabolism were the main metabolic pathways in feces and serum. A significant correlation between the gut microbiota and fecal metabolites was found (P < 0.01). Fecal metabolites were not greatly affected by the age or gender of giant pandas, but serum metabolites were significantly affected by age and gender. The majority of different metabolites caused by age were higher in serum of younger giant pandas, including fatty acids, lipids, metabolites of bile acids, and intermediate products of vitamin D3. The majority of different metabolites caused by gender included fatty acids, phosphatidylcholine (PC), phosphatidylserine (PS), and phosphatidylethanolamine (PE). A separate feeding diet should be considered according to different ages and genders of giant panda. Therefore, our results could provide helpful suggestions to further protect captive giant pandas.

PHARMACOKINETICS OF A SINGLE DOSE OF FLURALANER ADMINISTERED ORALLY TO AMERICAN BLACK BEARS (URSUS AMERICANUS).

Sarcoptic mange continues to impact free-ranging mammal populations, including the American black bear (Ursus americanus). Administration of a single oral dose of fluralaner may be a viable treatment option for captive and free-ranging black bears affected by mange. This novel ectoparasitic in the isoxazoline class acts as an inhibitor of γ-aminobutyric acid (GABA)-gated chloride channels and l-glutamate-gated chloride channels (GluCls) and is commercially available in the United States as a flea and tick preventative medication for domestic dogs and cats. Pharmacokinetic parameters of fluralaner were evaluated in clinically healthy American black bear cubs (n = 10) administered a single oral dose of fluralaner at a targeted minimum dose of 25 mg/kg. Blood was collected at 24 hr and 7, 14, 21, 28, 35, 42, 49, 56, 63, and 70 days, and harvested plasma was analyzed for drug concentration using high-performance liquid chromatography. The average half-life (Ke t1/2) was determined to be 4.9 days, which is shorter than that published in domestic dogs. It was estimated that the average drug withdrawal time is approximately 64-72 days in this species.

SERUM TRACE NUTRIENT VALUES IN FOUR CAPTIVE GIANT PANDAS (AILUROPODA MELANOLEUCA).

Trace nutrients have been shown to play important health roles in both domestic and wild animals, but there has been little investigation into their values and effects in giant pandas (Ailuropoda melanoleuca). The goal of this study was to obtain serum vitamin and mineral values in a group of healthy, captive giant pandas (n = 4). Twenty-seven samples were obtained from the group from 2008 to 2012. Serum analytes measured included retinol, α-tocopherol, hydroxyvitamin D, cobalt (Co), copper (Cu), iron (Fe), manganese (Mn), molybdenum (Mo), selenium (Se), and zinc (Zn). The diet was evaluated for protein, fat, fiber, Ca, P, magnesium (Mg), potassium (K), sodium (Na), Fe, Zn, Cu, Mn, and Mo. Comparison with other giant panda nutrient values were made. Further research with increased sample sizes and different populations are warranted.

Spring fasting behavior in a marine apex predator provides an index of ecosystem productivity.

The effects of declining Arctic sea ice on local ecosystem productivity are not well understood but have been shown to vary inter-specifically, spatially, and temporally. Because marine mammals occupy upper trophic levels in Arctic food webs, they may be useful indicators for understanding variation in ecosystem productivity. Polar bears (Ursus maritimus) are apex predators that primarily consume benthic and pelagic-feeding ice-associated seals. As such, their productivity integrates sea ice conditions and the ecosystem supporting them. Declining sea ice availability has been linked to negative population effects for polar bears but does not fully explain observed population changes. We examined relationships between spring foraging success of polar bears and sea ice conditions, prey productivity, and general patterns of ecosystem productivity in the Beaufort and Chukchi Seas (CSs). Fasting status (≥7 days) was estimated using serum urea and creatinine levels of 1,448 samples collected from 1,177 adult and subadult bears across three subpopulations. Fasting increased in the Beaufort Sea between 1983-1999 and 2000-2016 and was related to an index of ringed seal body condition. This change was concurrent with declines in body condition of polar bears and observed changes in the diet, condition and/or reproduction of four other vertebrate consumers within the food chain. In contrast, fasting declined in CS polar bears between periods and was less common than in the two Beaufort Sea subpopulations consistent with studies demonstrating higher primary productivity and maintenance or improved body condition in polar bears, ringed seals, and bearded seals despite recent sea ice loss in this region. Consistency between regional and temporal variation in spring polar bear fasting and food web productivity suggests that polar bears may be a useful indicator species. Furthermore, our results suggest that spatial and temporal ecological variation is important in affecting upper trophic-level productivity in these marine ecosystems.

Effects of biometrics, location and persistent organic pollutants on blood clinical-chemical parameters in polar bears (Ursus maritimus) from Svalbard, Norway.

In the present study, blood clinical-chemical parameters (BCCPs) were analysed in 20 female and 18 male Svalbard polar bears (Ursus maritimus) captured in spring 2007. The aim was to study how age, body condition (BC), biometrics, plasma lipid content and geographical location may confound the relationship between persistent organic pollutants (POPs) including PCBs, HCB, chlordanes, DDTs, HCHs, mirex and OH-PCBs and the concentrations of 12 specific BCCPs (hematocrit [HCT], hemoglobin [HB], aspartate aminotransferase [ASAT], alanine aminotransferase [ALAT], γ-glutamyltransferase [GGT], creatine kinase [CK], triglycerides [TG], cholesterol [CHOL], high-density lipoprotein [HDL], creatinine (CREA], urea, potassium (K]), and to investigate if any of these BCCPs may be applied as potential biomarkers for POP exposure in polar bears. Initial PCA and O-PLS modelling showed that age, lipids, BC and geographical location (longitude and latitude) were important parameters explaining BCCPs in females. Following subsequent partial correlation analyses correcting for age and lipids, multiple POPs in females were still significantly correlated with HCT and HDL (all p < 0.05). In males, age, BM, BC and longitude were important parameters explaining BCCPs. Following partial correlation analyses correcting for age, biometrics, lipids and longitude in males, multiple POPs were significantly correlated with HCT, ASAT, GGT and CHOL (all p < 0.05). In conclusion, several confounding parameters has to be taken into account when studying the relations between BCCPs and POPs in polar bears. When correcting for these, in particular HCT may be used as a simple cost-efficient biomarker of POP exposure in polar bears. Furthermore, decreasing HDL concentrations and increasing CHOL concentration with increasing POP concentrations may indicate responses related to increased risk of cardiovascular disease. We therefore suggest to further study POP exposure and lipidome response to increase knowledge of the risk of cardiometabolic syndrome in polar bears.

Blood Lead Levels in Captive Giant Pandas.

Fifteen giant pandas (Ailuropoda melanoleuca) from the Chinese Conservation and Research Center for the Giant Panda (CCRCGP) in Bifengxia, Sichuan, China were analyzed for blood lead concentrations (Pb-B) during the 2017 breeding season. Thirteen of the 15 bears showed Pb-B below the method detection limit (MDL) of 3.3 µg/dL. The two remaining bears, although above the MDL, contained very low concentrations of lead of 3.9 and 4.5 µg/dL. All 15 giant pandas in this analysis had Pb-B concentrations that were within normal background concentrations for mammals in uncontaminated environments. For a threatened species, whose native country is plagued by reports of extremely high air pollution, our findings suggest that giant pandas at the CCRCGP are not absorbing lead at concentrations that would adversely affect their health.

SURVEY FOR EQUINE HERPESVIRUSES IN POLAR BEARS ( URSUS MARITIMUS) AND EXOTIC EQUIDS HOUSED IN US AZA INSTITUTIONS.

Infection by equine herpesvirus (EHV) strains (EHV-1, EHV-9) in ursid species, including polar bears ( Ursus maritimus), has been associated with neurological disease and death. A serosurvey of captive exotic equid and polar bear populations in US Association of Zoos and Aquaria institutions was performed to determine the prevalence of EHV strains using quantitative polymerase chain reaction (qPCR) and enzyme-linked immunosorbent assay (ELISA) tests. Equid species surveyed included zebra ( Equus spp.), Przewalski's wild horse ( Equus ferus przewalskii), Persian onager ( Equus hemionus), and Somali wild ass ( Equus africanus somaliensis). A questionnaire regarding husbandry and medical variables was distributed to institutions housing polar bears. No polar bears tested positive for EHVs on qPCR of blood or nasal swabs. No exotic equids tested positive for EHVs on qPCR of blood, but two exotic equids ( n = 2/22; 9%) tested positive for EHVs on qPCR of nasal swabs. On ELISA, polar bears infrequently were positive for EHV-1 ( n = 5/38; 13%). Exotic equids were positive for EHV-4 on ELISA more frequently ( n = 30/43; 70%) than for EHV-1 ( n = 8/43; 19%). Nine institutions submitted samples from both exotic equids and polar bears, two of which had both exotic equids and polar bears positive for EHVs by ELISA. Each of these institutions reported that the polar bear and exotic equid exhibits were within 80 m of each other and that risk factors for fomite transmission between exhibits based on husbandry practices were present. One institution that did not house exotic equids had a polar bear test positive for EHV-1 on ELISA, with no history of exposure to exotic equids. Further testing of captive polar bears and exotic equids is recommended, as is modification of husbandry practices to limit exposure of polar bears to exotic equids.

Isotopic Composition of Blood of Polar Bears (Ursus maritimus) of the Kara-Barents Sea Population.

The data on the content of carbon and nitrogen isotopes in the blood samples of polar bears obtained in the present study confirm that polar bears in the Taimyr region (and the Kara-Barents sea population in general) are partly dependent on the resources of terrestrial origin. However the "terrestrial carbon" evidently reaches bears' tissues indirectly, via marine food webs utilizing organic carbon brought into the polar basin by Siberian rivers.

Hundreds of Unrecognized Halogenated Contaminants Discovered in Polar Bear Serum.

Exposure of polar bears (Ursus maritimus) to persistent organic pollutants was discovered in the 1970s, but recent evidence suggests the presence of unknown toxic chemicals in their blood. Protein and phospholipid depleted serum was stirred with polyethersulfone capillaries to extract a broad range of analytes, and nontarget mass spectrometry with "fragmentation flagging" was used for detection. Hundreds of analytes were discovered belonging to 13 classes, including novel polychlorinated biphenyl (PCB) metabolites and many fluorinated or chlorinated substances not previously detected. All analytes were detected in the oldest (mid-1980s) archived polar bear serum from Hudson Bay and Beaufort Sea, and all fluorinated classes showed increasing trends. A mouse experiment confirmed the novel PCB metabolites, suggesting that these could be widespread in mammals. Historical exposure and toxic risk has been underestimated, and these halogenated contaminants pose uncertain risks to this threatened species.

Blood clotting behavior is innately modulated in Ursus americanus during early and late denning relative to summer months.

Remarkably, American black bears (Ursus americanus) are capable of varying their heart rates to coincide with their breathing, creating pauses of 30 s or more, yet they do not appear to suffer from embolic events. We evaluated some features of the clotting cascade of black bears, providing novel insights into the underlying mechanisms they evoke for embolic protection during hibernation. We measured activated clotting time, prothrombin time and activated partial thromboplastin time during early denning (December), late denning (March) and summer (August). Activated clotting time during early hibernation was ∼3 times longer than that observed among non-hibernating animals. Clotting time was reduced later in hibernation, when bears were within ∼1 month of emerging from dens. Prothrombin time was similar for each seasonal time point, whereas activated partial thromboplastin time was highest during early denning and decreased during late denning and summer. We also examined D-dimer concentration to assess whether the bears were likely to have experienced embolic events. None of the non-parturient bears exceeded a D-dimer concentration of 250 ng ml-1 (considered the clinical threshold for embolism in mammals). Our findings suggest there is unique expression of the clotting cascade in American black bears during hibernation, in which extrinsic pathways are maintained but intrinsic pathways are suppressed. This was evaluated by a significant difference between the activated clotting time and activated partial thromboplastin time during the denning and non-denning periods. These changes are likely adaptive, to avoid clotting events during states of immobilization and/or periods of asystole. However, an intact extrinsic pathway allows for healing of external injuries and/or foreign body responses.

HEMATOLOGY, SERUM BIOCHEMISTRY, AND URINALYSIS VALUES IN THE ADULT GIANT PANDA ( AILUROPODA MELANOLEUCA).

The giant panda ( Ailuropoda melanoleuca) is a high-profile threatened species with individuals in captivity worldwide. As a result of advances in captive animal management and veterinary medicine, the ex situ giant panda population is aging, and improved understanding of age-related changes is necessary. Urine and blood samples were collected in April and July 2015 and analyzed for complete blood count, serum biochemistry, and biochemical and microscopic urine analysis for all individuals sampled ( n = 7, 7-16 yr of age) from giant panda housed at the China Research and Conservation Centre for the Giant Panda in Bifengxia, Sichuan Province, China. Hematology and serum biochemistry values were similar to those previously reported for giant panda aged 2-20 yr and to Species360 (formerly International Species Information System) values. Urine was overall dilute (urine specific gravity range: 1.001-1.021), acellular, and acidic (pH range: 6-7). This is the first report of hematologic and serum biochemistry, with associated urinalysis values, in the giant panda aged 7-16 yr.

ANTIDOG IgG SECONDARY ANTIBODY SUCCESSFULLY DETECTS IgG IN A VARIETY OF AQUATIC MAMMALS.

Serological tests play an important role in the detection of wildlife diseases. However, while there are many commercial assays and reagents available for domestic species, there is a need to develop efficient serological assays for wildlife. In recent years, marine mammals have represented a wildlife group with emerging infectious diseases, such as influenza, brucellosis, and leptospirosis. However, with the exception of disease-agent-specific assays or functional assays, few reports describe the use of antibody detection assays in marine mammals. In an indirect enzyme-linked immunoassay (EIA) or an immunofluorescence assay, antibody is detected using an antitarget species secondary conjugated antibody. The sensitivity of the assay depends on the avidity of the binding reaction between the bound antibody and the detection antibody. A commercial polyclonal antidog IgG conjugated antibody was tested in an EIA for its ability to sensitively detect the IgG of seven marine mammals including sea otter ( Enhydra lutris ), polar bear ( Ursus maritimus ), grey seal ( Halichoerus grypus ), harbor seal ( Phoca vitulina ), northern elephant seal ( Mirounga angustirostris ), California sea lion ( Zalophus californianus ), Pacific walrus ( Odobenus rosmarus ) and one freshwater mammal: Asian small-clawed otter ( Aonyx cinerea ). With the exception of Asian small-clawed sea otters, the detection of IgG in these marine mammals either exceeded or was nearly equal to detection of dog IgG. The use of the tested commercial antidog IgG antibody may be a valid approach to the detection of antibody response to disease in sea mammals.

Suppressed bone remodeling in black bears conserves energy and bone mass during hibernation.

Decreased physical activity in mammals increases bone turnover and uncouples bone formation from bone resorption, leading to hypercalcemia, hypercalcuria, bone loss and increased fracture risk. Black bears, however, are physically inactive for up to 6 months annually during hibernation without losing cortical or trabecular bone mass. Bears have been shown to preserve trabecular bone volume and architectural parameters and cortical bone strength, porosity and geometrical properties during hibernation. The mechanisms that prevent disuse osteoporosis in bears are unclear as previous studies using histological and serum markers of bone remodeling show conflicting results. However, previous studies used serum markers of bone remodeling that are known to accumulate with decreased renal function, which bears have during hibernation. Therefore, we measured serum bone remodeling markers (BSALP and TRACP) that do not accumulate with decreased renal function, in addition to the concentrations of serum calcium and hormones involved in regulating bone remodeling in hibernating and active bears. Bone resorption and formation markers were decreased during hibernation compared with when bears were physically active, and these findings were supported by histomorphometric analyses of bone biopsies. The serum concentration of cocaine and amphetamine regulated transcript (CART), a hormone known to reduce bone resorption, was 15-fold higher during hibernation. Serum calcium concentration was unchanged between hibernation and non-hibernation seasons. Suppressed and balanced bone resorption and formation in hibernating bears contributes to energy conservation, eucalcemia and the preservation of bone mass and strength, allowing bears to survive prolonged periods of extreme environmental conditions, nutritional deprivation and anuria.

Seasonal variation in haematological and biochemical variables in free-ranging subadult brown bears (Ursus arctos) in Sweden.

BACKGROUND: Free-ranging brown bears exhibit highly contrasting physiological states throughout the year. They hibernate 6 months of the year, experiencing a decrease in body temperature, heart rate, respiratory rate and metabolism. An increase in food consumption and the resulting weight gain (mostly through fat storage) prior to hibernation are also part of the brown bear's annual cycle. Due to these physiological changes, haematological and biochemical variables vary dramatically throughout the year. Seasonal changes in 12 haematological and 34 biochemical variables were evaluated in blood samples collected from 40 free-ranging subadult brown bears (22 females, 18 males) immobilised in Sweden in winter (February-March), spring (April-May), and summer (June). RESULTS: Higher levels of haemoglobin, haematocrit and red blood cell count, and a lower white blood cell count and mean cell volume was found during hibernation than in spring and summer. Lower values of the enzymes; aspartate aminotransferase (AST), alanine transaminase (ALT), alkaline phosphatase (AP), γ-glutamyl transpeptidase (GGT), glutamate dehydrogenase (GD) and amylase, and increased values of ß-hydroxybutyrate (ß-HBA) and blood lipids; triglycerides, cholesterol and free fatty acids, were present during hibernation compared to spring and summer. CONCLUSIONS: This study documents significant shifts in haematological and biochemical variables in samples collected from brown bears anaesthetised in winter (February-March) compared to in spring and summer (April-June), reflecting the lowered metabolic, renal and hepatic activity during hibernation. Lower values of enzymes and higher values of blood lipids during hibernation, likely reflect a lipid-based metabolism.

Identification of conserved microRNAs in peripheral blood from giant panda: expression of mammary gland-related microRNAs during late pregnancy and early lactation.

The giant panda (Ailuropoda melanoleuca) is one of the world's most endangered mammals, and it has evolved several unusual biological and behavioral traits. During puberty, pregnancy, lactation, and involution, the mammary gland undergoes profound morphological and functional changes. A large number of microRNAs (miRNAs) have been identified to be involved in mammary gland development and lactation. In this study, we identified 202 conserved mature miRNAs, corresponding to 147 pre-miRNAs, in giant panda peripheral blood using a small RNA-sequencing approach. In addition, 27 miRNA families and 29 miRNA clusters were identified. We analyzed the arm selection preference of pre-miRNAs and found that: 1) most giant panda pre-miRNAs generated one-strand miRNAs, and the 5p-arm only miRNAs have a higher expression level than 3p-arm only miRNAs; 2) there were more 5p-arm dominant miRNAs than 3p-arm dominant miRNAs; and 3) 5p-arm dominant miRNAs have a larger fold change within miRNA pairs than 3p-arm dominant miRNAs. Expression of 12 lactation-related miRNAs was detected across late pregnancy and early lactation stages by qPCR, and seven miRNAs were identified as clustered in one significant model. Most of these clustered miRNAs exhibited inhibitory roles in proliferation and differentiation of mammary epithelial cells. Functional analysis highlighted important roles of the seven as signed miRNAs in mammary development and metabolic changes, including blood vessel morphogenesis, macromolecule biosynthesis, cell cycle regulation, and protein transport.

Winter blood values of selected parameters in a group of non-hibernating captive brown bears (Ursus arctos).

Bears undergo some significant changes reflected in blood values during winter season. The most significant are reduced urea and increased creatinine, by some authors considered to be physiological indicators of hibernation. Studied group of six captive brown bears (Ursus arctos) showed decreased activity in winter but were accepting food and walked outdoors. Blood parameters assessed in February 2011 revealed mean values of leucocytes and neutrophils as significantly lower, and creatinine significantly increased compared to captive and free living bears sampled during other seasons when bears are active.