A Novel Approach to Identifying Physical Markers of Cryo-Damage in Bull Spermatozoa

Cryopreservation is an efficient way to store spermatozoa and plays a critical role in the livestock industry as well as in clinical practice. During cryopreservation, cryo-stress causes substantial damage to spermatozoa. In present study, the effects of cryo-stress at various cryopreservation steps, such as dilution / cooling, adding cryoprtectant, and freezing were studied in spermatozoa collected from 9 individual bull testes. The motility (%), motion kinematics, capacitation status, mitochondrial activity, and viability of bovine spermatozoa at each step of the cryopreservation process were assessed using computer-assisted sperm analysis, Hoechst 33258/chlortetracycline fluorescence, rhodamine 123 staining, and hypo-osmotic swelling test, respectively. The results demonstrate that the cryopreservation steps reduced motility (%), rapid speed (%), and mitochondrial activity, whereas medium/slow speed (%), and the acrosome reaction were increased (P < 0.05). Differences (Δ) of the acrosome reaction were higher in dilution/cooling step (P < 0.05), whereas differences (Δ) of motility, rapid speed, and non-progressive motility were higher in cryoprotectant and freezing as compared to dilution/cooling (P < 0.05). On the other hand, differences (Δ) of mitochondrial activity, viability, and progressive motility were higher in freezing step (P < 0.05) while the difference (Δ) of the acrosome reaction was higher in dilution/cooling (P < 0.05). Based on these results, we propose that freezing / thawing steps are the most critical in cryopreservation and may provide a logical ground of understanding on the cryo-damage. Moreover, these sperm parameters might be used as physical markers of sperm cryo-damage.     

Molecular Survey of Bacterial Communities Associated with Bacterial Chondronecrosis with Osteomyelitis (BCO) in Broilers

Bacterial chondronecrosis with osteomyelitis (BCO) is recognized as an important cause of lameness in commercial broiler chickens (meat-type chickens). Relatively little is known about the microbial communities associated with BCO. This study was conducted to increase our understanding of the microbial factors associated with BCO using a culture-independent approach. Using Illumina sequencing of the hyper-variable region V6 in the 16S rRNA gene, we characterized the bacterial communities in 97 femoral or tibial heads from normal and lame broilers carefully selected to represent diverse variations in age, line, lesion type, floor type, clinical status and bone type. Our in-depth survey based on 14 million assembled sequence reads revealed that complex bacterial communities exist in all samples, including macroscopically normal bones from clinically healthy birds. Overall, Proteobacteria (mean 90.9%) comprised the most common phylum, followed by Firmicutes (6.1%) and Actinobacteria (2.6%), accounting for more than 99% of all reads. Statistical analyses demonstrated that there are differences in bacterial communities in different types of bones (femur vs. tibia), lesion types (macroscopically normal femora or tibiae vs. those with pathognomonic BCO lesions), and among individual birds. This analysis also showed that BCO samples overrepresented genera Staphylococcus, whose species have been frequently isolated in BCO samples in previous studies. Rarefaction analysis demonstrated the general tendency that increased severities of BCO lesions were associated with reduced species diversity in both femoral and tibial samples when compared to macroscopically normal samples. These observations suggest that certain bacterial subgroups are preferentially selected in association with the development of BCO lesions. Understanding the microbial species associated with BCO will identify opportunities for understanding and modulating the pathogenesis of this form of lameness in broilers.     

Insulin Resistance Is Associated with Prevalence of Physician-Diagnosed Urinary Incontinence in Postmenopausal Non-Diabetic Adult Women: Data from the Fourth Korea National Health and Nutrition Examination Survey

Objective

To investigate the association between insulin resistance (IR) and urinary incontinence in Korean adult women by analyzing the data from the Korea National Health and Nutrition Examination Survey IV (KNHANES) 2007–2009

Methods

A nationally representative sample of 5318 non-diabetic Korean women ≥19-years-of-age (3043 premenopausal and 2275 postmenopausal women) was included from KNHANES 2008–2010. IR was measured using the homeostasis model assessment of IR (HOMA-IR). Participants in the highest and lowest quartile of HOMA-IR were defined as insulin-resistant and insulin-sensitive respectively. Women who have current physician-diagnosed urinary incontinence were classified as having urinary incontinence.

Results

Incontinence was found in 9.18% of the total population, 8.51% of the premenopausal population, and 10.86% of the postmenopausal population. The prevalence of incontinence increased with age, reaching a peak at 60-69-years-of-age. The prevalence of urinary incontinence increased significantly with higher HOMA-IR quartiles in pre- and post-menopausal women (p for linear association = 0.0458 and 0.0009 respectively). Among post-menopausal women, those in the highest quartile of HOMA-IR were significantly more likely to have urinary incontinence compared to those in the lowest quartile [adjusted odds ratio, 1.72; 95% confidence interval, 1.07–2.77]. However premenopausal population exhibited no association between incontinence and HOMA-IR quartiles

Conclusion

Our results suggest that the prevalence of incontinence increased across HOMA-IR in non-diabetic adult women, and especially, IR might be a risk factor for incontinence in postmenopausal non-diabetic women.     

Protective effects of PEP-1-Catalase on stress-induced cellular toxicity and MPTP-induced Parkinson’s disease

Parkinson’s disease (PD) is a neurodegenerative disability caused by a decrease of dopaminergic neurons in the substantia nigra (SN). Although the etiology of PD is not clear, oxidative stress is believed to lead to PD. Catalase is antioxidant enzyme which plays an active role in cells as a reactive oxygen species (ROS) scavenger. Thus, we investigated whether PEP-1-Catalase protects against 1-methyl-4-phenylpyridinium (MPP⁺) induced SH-SY5Y neuronal cell death and in a 1-methyl-4-phenyl-1,2,3,6-trtrahydropyridine (MPTP) induced PD animal model. PEP-1-Catalase transduced into SH-SY5Y cells significantly protecting them against MPP⁺-induced death by decreasing ROS and regulating cellular survival signals including Akt, Bax, Bcl-2, and p38. Immunohistochemical analysis showed that transduced PEP-1-Catalase markedly protected against neuronal cell death in the SN in the PD animal model. Our results indicate that PEP-1-Catalase may have potential as a therapeutic agent for PD and other oxidative stress related diseases. [BMB Reports 2015; 48(7): 395-400]     

Simple Modification of the Bladder Outlet Obstruction Index for Better Prediction of Endoscopically-Proven Prostatic Obstruction: A Preliminary Study

Purpose

The bladder outlet obstruction index (BOOI), also known as the Abrams-Griffiths (AG) number, is the most widely used index for predicting BOO. However, the obstructed prostatic urethra determined by the BOOI is often inconsistent with endoscopically-proven obstruction. We assessed abdominal straining pattern as a novel parameter for improving the prediction of BOO.

Materials and Methods

We retrospectively reviewed the pressure-flow studies (PFS) and cystourethroscopy in 176 BPH/LUTS patients who were unresponsive to medical therapy. During PFS, some groups of patients tried to urinate with abdominal straining, which can increases intravesical pressure and underestimate BOOI theoretically. Accordingly, the modified BOOI was defined as (P_detQ_max+ΔP_abd)-2Q_max.

Results

Ultimately, 130 patients were eligible for the analysis. In PFS, ΔP_abd (P_abdQ_max-initial P_abd) was 11.81±13.04 cmH₂O, and it was 0–9 cmH₂O in 75 (57.7%), 10–19 cmH₂O in 23 (17.7%) and ≥20 cmH₂O in 32 (24.6%) patients. An endoscopically obstructed prostatic urethra in 92 patients was correctly determined in 47 patients (51.1%) by the original BOOI versus 72 patients (78.3%) based on the modified BOOI. Meanwhile, an “unobstructed” urethra according to the original BOOI was present in 11 patients (12.0%), whereas according to the modified BOOI, only 2 (2.1%) would be labeled as “unobstructed”. In receiver operating characteristic curves, the area under the curve was 0.906 using the modified BOOI number versus 0.849 in the original BOOI (p<0.05).

Conclusions

The change in abdominal pressure was correlated with endoscopically-proven obstruction. Our simple modification of the BOOI on the basis of this finding better predicted bladder outlet obstruction and, therefore, should be considered when evaluating BOO in patients with LUTS/BPH.     

Chronic Obstructive Pulmonary Disease-Related Non-Small-Cell Lung Cancer Exhibits a Low Prevalence of EGFR and ALK Driver Mutations

Lung cancer and chronic obstructive pulmonary disease (COPD) are two major lung diseases. Epidermal growth factor receptor (EGFR) mutations, v‐Ki‐ras2 Kirsten rat sarcoma (KRAS) mutations and anaplastic lymphoma kinase (ALK) gene rearrangements represent driver mutations that are frequently assessed on initial evaluation of non-small-cell lung cancer (NSCLC). The present study focused on the expression of driver mutations in NSCLC patients presenting with COPD and further evaluated the association between NSCLC and COPD. Data from 501 consecutive patients with histologically proven recurrent or metastatic NSCLC were analyzed retrospectively. The patients underwent spirometry and genotyping of EGFR, ALK, and KRAS in tissue samples. Patient characteristics and expression of driver mutations were compared between the COPD and non-COPD groups.Among 350 patients with spirometric results, 106 (30.3%) were diagnosed with COPD, 108 (30.9%) had EGFR mutations, 31 (8.9%) had KRAS mutations, and 34 (9.7%) showed ALK rearrangements. COPD was independently associated with lower prevalences of EGFR mutations (95% confidence interval [CI], 0.254–0.931, p = 0.029) and ALK rearrangements (95% CI, 0.065–0.600, p = 0.004). The proportions of EGFR mutations and ALK rearrangements decreased as the severity of airflow obstruction increased (p = 0.001). In never smokers, the prevalence of EGFR mutations was significantly lower in the COPD group than in the non-COPD group (12.7% vs. 49.0%, p = 0.002). COPD-related NSCLC patients exhibited low prevalences of EGFR mutations and ALK rearrangements compared with the non-COPD group. Further studies are required regarding the molecular mechanisms underlying lung cancer associated with COPD.     

Serum levels of appetite-regulating hormones and pro-inflammatory cytokines are ameliorated by a CLA diet and endurance exercise in rats fed a high-fat diet

[Purpose]

This study examined whether conjugated linoleic acid (CLA) supplementation and endurance exercise affect appetite-regulating hormones and pro-inflammatory cytokines in rats.

[Methods]

Seven-week-old male Sprague-Dawley rats were divided randomly into the high-fat diet sedentary group (HS, n=8), the 1.0% CLA supplemented high-fat diet sedentary group (CS, n=8), and the 1.0% CLA supplemented high-fat diet exercise group (CE, n=8). Rats in the CE group swam 60 min/day, 5 days/week for 4 weeks.

[Results]

Leptin and insulin levels in the CS and CE groups were significantly lower than those in the HS group (p<0.001), whereas leptin (p<0.01) and insulin (p<0.05) levels decreased significantly in the CE compared to those in the CS group. Interleukin (IL)-1β (p<0.001) and IL-6 (p<0.01) levels in the CS and CE groups decreased significantly compared to those in the HS group. Leptin (IL-1β: r=0.835, p<0.001), IL-6 (r=0.607, p<0.05), insulin (IL-1β: r=0.797, p<0.01), and IL-6 (r=0.827, p<0.01) levels were positively related with pro-inflammatory cytokine levels.

[Conclusion]

Endurance exercise may play an important role during CLA supplementation of rats on a high-fat diet.     

The change in peripheral sweating mechanisms of the tropical Malaysian who stays in Japan

Tropical subjects regulate core temperature with less amount of sweat against heat compared to temperate subjects through long-term heat-acclimatization. The purpose of the study is to determine whether acclimatization in tropical subjects decay during a stay in temperate area. The aim of this study, therefore, was to investigate the possible changes in the peripheral sweating mechanisms. Local sweating response activated by acetylcholine (ACh) applied iontophoretically among Malaysians with varying duration of stay in Japan and Japanese resident subjects. Directly activated (DIR) and axon reflex (AXR)-mediated sweating during ACh iontophoresis were measured by capacitance hygrometer (quantitative sudomotor axon reflex test, QSART) QSART was performed in a thermoneutral condition (24±0.5 °C, 40±3% rh). The sweat onset-time after the current loading was 1.05 min shorter in Malaysian with long-term stay in Japan (MLJ) than in Malaysian, and the AXR(1), AXR(2) and DIR sweating in MLJ were larger than Malaysian. From these results, suppressed neuroglandular response to ACh was confirmed in Malaysians. It is suggested that long-term heat-acclimatization acquired in tropical subjects may decay after immigration to temperate area.     

Transgenic nonhuman primates for neurodegenerative diseases

Animal models that represent human diseases constitute an important tool in understanding the pathogenesis of the diseases, and in developing effective therapies. Neurodegenerative diseases are complex disorders involving neuropathologic and psychiatric alterations. Although transgenic and knock-in mouse models of Alzheimer's disease, (AD), Parkinson's disease (PD) and Huntington's disease (HD) have been created, limited representation in clinical aspects has been recognized and the rodent models lack true neurodegeneration. Chemical induction of HD and PD in nonhuman primates (NHP) has been reported, however, the role of intrinsic genetic factors in the development of the diseases is indeterminable. Nonhuman primates closely parallel humans with regard to genetic, neuroanatomic, and cognitive/behavioral characteristics. Accordingly, the development of NHP models for neurodegenerative diseases holds greater promise for success in the discovery of diagnoses, treatments, and cures than approaches using other animal species. Therefore, a transgenic NHP carrying a mutant gene similar to that of patients will help to clarify our understanding of disease onset and progression. Additionally, monitoring disease onset and development in the transgenic NHP by high resolution brain imaging technology such as MRI, and behavioral and cognitive testing can all be carried out simultaneously in the NHP but not in other animal models. Moreover, because of the similarity in motor repertoire between NHPs and humans, it will also be possible to compare the neurologic syndrome observed in the NHP model to that in patients. Understanding the correlation between genetic defects and physiologic changes (e.g. oxidative damage) will lead to a better understanding of disease progression and the development of patient treatments, medications and preventive approaches for high risk individuals. The impact of the transgenic NHP model in understanding the role which genetic disorders play in the development of efficacious interventions and medications is foreseeable.