The significance of growth shells in development of symmetry, transparency, and refraction of the human lens.

Human visual function depends on the biological lens, a biconvex optical element formed by coordinated, synchronous generation of growth shells produced from ordered cells at the lens equator, the distal edge of the epithelium. Growth shells are comprised of straight (St) and S-shaped (SSh) lens fibers organized in highly symmetric, sinusoidal pattern which optimizes both the refractile, transparent structure and the unique microcirculation that regulates hydration and nutrition over the lifetime of an individual. The fiber cells are characterized by diversity in composition and age. All fiber cells remain interconnected in their growth shells throughout the life of the adult lens. As an optical element, cellular differentiation is constrained by the physical properties of light and its special development accounts for its characteristic symmetry, gradient of refractive index (GRIN), short range transparent order (SRO), and functional longevity. The complex sinusoidal structure is the basis for the lens microcirculation required for the establishment and maintenance of image formation.

Cross-jurisdictional pandemic management: providers speaking on the experience of Nunavut Inuit accessing services in Manitoba during the COVID-19 pandemic.

Across Canada, the COVID-19 pandemic placed considerable stress on territorial and provincial healthcare systems. For Nunavut, the need to continue to provide access to critical care to its citizens meant that medical travel to provincial points of care (Edmonton, Winnipeg and Ottawa) had to continue through the pandemic. This complexity created challenges related to the need to keep Nunavut residents safe while accessing care, and to manage the risk of outbreaks in Nunavut resultant from patients returning home. A number of strategies were adopted to mitigate risk, including the expansion of virtual care, self-isolation requirements before returning from Winnipeg, and a level of cross-jurisdictional coordination previously unprecedented. Structural limitations in Nunavut however limited opportunities to expand virtual care, and to allow providers from Manitoba to access the Nunavut's electronic medical records of patients requiring follow up. Thus, known and long-standing issues exacerbated vulnerabilities within the Nunavut healthcare system. We conclude that addressing cross-jurisdictional issues would be well served by the development of a more formal Nunavut-Manitoba agreement (with similar agreements with Ontario and Alberta), outlining mutual obligations and accountabilities.

Mitigating the impact of the COVID-19 pandemic on Inuit living in Manitoba: community responses.

We document community responses to the COVID-19 pandemic among Inuit living in the province of Manitoba, Canada. This study was conducted by the Manitoba Inuit Association and a Council of Inuit Elders, in partnership with researchers from the University of Manitoba. We present findings from 12 health services providers and decision-makers, collected in 2021.Although Public Health orders led to the closure of the Manitoba Inuit Association's doors to community events and drop-in activities, it also created opportunities for the creation of programming and events delivered virtually and through outreach. The pandemic exacerbated pre-existing health and social system's shortcomings (limited access to safe housing, food insecurity) and trauma-related tensions within the community. The Manitoba Inuit Association achieved unprecedented visibility with the provincial government, receiving bi-weekly reports of COVID-19 testing, results and vaccination rates for Inuit. We conclude that after over a decade of advocacy received with at best tepid enthusiasm by federal and provincial governments, the Manitoba Inuit Association was able effectively advocate for Inuit-centric programming, and respond to Inuit community's needs, bringing visibility to a community that had until then been largely invisible. Still, many programs have been fueled with COVID-19 funding, raising the issue of sustainability.

We must take a One Health approach to improve pandemic infection control.

The Covid-19 pandemic must serve as a wake-up call to work more collaboratively between medical and veterinary practitioners, biologists and environmentalists say Camilla Benfield, David Heymann, Judy MacArthur Clark, AJ Trees and Babulal Sethia.

Guidelines for the ethical review of laboratory animal welfare People's Republic of China National Standard GB/T 35892-2018 [Issued 6 February 2018 Effective from 1 September 2018].

These Chinese National Guidelines (GB/T 35892-20181) were issued February 06, 2018 and became effective September 01, 2018. The authors recognized the urgent need for an authentic English translation to inform the international community of the compliance requirements in China. It was appreciated that the final translation must reflect the specialist understanding of those working under the Guideline whilst remaining faithful to the meaning of the original Chinese text. A three-step translation process was therefore determined. Step 1: A professional interpretation service (KL Communications, UK) was commissioned to prepare a literal translation of the Chinese text. Supportive documents were provided which explained specialist terminology. This translation was checked by two bilingual experts. Step 2: A workshop was held in Nanjing in May 2019 to which were invited experts in laboratory animal welfare and ethical use. These included international native English-speaking and Chinese-speaking delegates. The delegates worked in multi-lingual teams to review sections of the literal translation ahead of the workshop, and to agree an authentic interpretation during the workshop. Step 3: Following the workshop, three bilingual experts (two native Chinese speakers and one native English speaker) reviewed the entire document to ensure consistency of terminology and general accuracy. This document is thus not a "literal translation" but an "accurate interpretation" of the original text. Any challenge of work being performed under these Guidelines should rely on the Chinese text in the first place. However, this translation may be used as mitigating evidence, especially where those performing the work are non-Chinese speakers.

Communicating About Animal Research with the Public.

Animals play a key role in biomedical research and other areas of scientific inquiry. But public opinion plays a key role in influencing how this area of science is regulated and funded. Nevertheless, scientists have historically been reticent to speak openly about their animal research or to open their animal facilities to the public in any way. Consequently, most of the available information has come from those opposed to animal research. This imbalance has led to suspicion and lagging public support for this work. To reverse this effect, efforts are now being made in many parts of the world to increase openness and transparency in this sector. The authors firmly believe that encouraging more institutions to join this movement, focused on better and greater communication, is essential to preserve the research community's "permission" to perform justifiable studies involving animals. For the purposes of this article, we consider "the public" to include that cross-section of society who may be asked their views in opinion poll studies and who may vote in elections. It also includes other influential groups such as the media, scientists working in other disciplines, animal welfare groups, and politicians who may shape regulatory frameworks. Public opinion on this issue matters. The majority of funding for biomedical research comes, either directly or indirectly, from the public purse. In the case of pharmaceutical research, funding derives from selling medicines to consumers. We therefore all have a vested interest in this funding. Furthermore, legislation that covers the use of animals in research is permissive-it allows scientists to do things that might otherwise contravene animal welfare laws. But this permission is normally contingent on complying with strict protective measures designed to ensure the work stays within the ethical framework that public opinion has deemed appropriate. Open and transparent communication is the best way to promote public understanding. There is thus a responsibility on all those involved in animal research, whether scientists, animal care staff, physicians, veterinarians, members of ethics committees, or managers and leaders, to support and promote public awareness and trust in this work. Circumstantial evidence shows that, with such open dialogue, there is decreased targeting and harassment of individuals and job pride and satisfaction for all involved is improved.

The 3Rs in research: a contemporary approach to replacement, reduction and refinement.

First promulgated in 1959, the 3Rs of Replacement, Reduction and Refinement have evolved as fundamental principles underlying the use of animals and alternatives in science throughout the modern world. This review describes a contemporary approach to delivering the 3Rs through acknowledging the contribution of new technologies and emphasising that applying the 3Rs can be beneficial to good science as well as to animal welfare. This science-led approach moves the concept of the 3Rs out of an ethical silo where they were often considered by scientists to be an inconvenient obligation. On the contrary, relevant examples demonstrate the opportunity to practise better science using 3Rs technologies which deliver faster, more reproducible and more cost-effective results. Indeed, methods harnessing Replacement approaches may permit discoveries which are simply not feasible using animals and frequently are more flexible and agile since compliance with regulatory oversight requirements is simplified. Although the necessity for rigorous oversight is well recognised, it is important that the associated bureaucracy is not allowed to become prohibitive, causing scientists to avoid pursuing justifiable and important research involving animals. Public support for research is conditional - animals should not suffer unnecessarily and sufficient potential benefit should accrue from the research. However, society also actively seeks pioneering medical and scientific advances which can only be achieved through research. Therefore, a balance must be struck between safeguarding animal welfare whilst enabling high-quality science. It is this balance which promotes and sustains public confidence that animal based research is acceptable and being appropriately managed.

Roles of the International Council for Laboratory Animal Science (ICLAS) and International Association of Colleges of Laboratory Animal Medicine (IACLAM) in the Global Organization and Support of 3Rs Advances in Laboratory Animal Science.

Practical implementation of the 3Rs at national and regional levels around the world requires long-term commitment, backing, and coordinated efforts by international associations for laboratory animal medicine and science, including the International Association of Colleges of Laboratory Animal Medicine (IACLAM) and the International Council for Laboratory Animal Science (ICLAS). Together these organizations support the efforts of regional organization and communities of laboratory animal science professionals as well as the development of local associations and professional colleges that promote the training and continuing education of research facility personnel and veterinary specialists. The recent formation of a World Organization for Animal Health (OIE) Collaborating Center for Laboratory Animal Science and Welfare emphasizes the need for research into initiatives promoting laboratory animal welfare, particularly in emerging economies and regions with nascent associations of laboratory animal science.

A method to prevent protein delocalization in imaging mass spectrometry of non-adherent tissues: application to small vertebrate lens imaging.

MALDI imaging requires careful sample preparation to obtain reliable, high-quality images of small molecules, peptides, lipids, and proteins across tissue sections. Poor crystal formation, delocalization of analytes, and inadequate tissue adherence can affect the quality, reliability, and spatial resolution of MALDI images. We report a comparison of tissue mounting and washing methods that resulted in an optimized method using conductive carbon substrates that avoids thaw mounting or washing steps, minimizes protein delocalization, and prevents tissue detachment from the target surface. Application of this method to image ocular lens proteins of small vertebrate eyes demonstrates the improved methodology for imaging abundant crystallin protein products. This method was demonstrated for tissue sections from rat, mouse, and zebrafish lenses resulting in good-quality MALDI images with little to no delocalization. The images indicate, for the first time in mouse and zebrafish, discrete localization of crystallin protein degradation products resulting in concentric rings of distinct protein contents that may be responsible for the refractive index gradient of vertebrate lenses.

Implementing the 3Rs in neuroscience research: a reasoned approach.

The 3Rs--replacement, reduction, and refinement--are aimed at minimizing the welfare costs to animals used in research. Some neuroscientists fear that implementing the 3Rs will prohibit essential studies. Others view them as fundamental ethical principles that improve the quality of research. A regulatory system that integrates science and welfare is most likely to deliver public confidence.

Development and application of a novel environmental preference chamber for assessing responses of laboratory mice to atmospheric ammonia.

A novel environmental preference chamber (EPC) was developed and used to assess responses of laboratory mice to atmospheric ammonia. The EPC features 1) a test chamber with 4 individually ventilated, mutually accessible compartments; b) automated tracking of mouse movements by using paired infrared sensors; c) identification of individual mice by using photosensors; d) monitoring and regulation of the NH3 concentration in each compartment; and e) personal-computer-based data acquisition. In an initial preference study with the EPC, 4 groups of 4 laboratory mice (BALB/c/Bkl; body weight, 13.4 to 18.4 g) were each given a choice among 4 NH3 concentrations (mean +/- SE) of 4 +/- 2, 30 +/- 2, 56 +/- 4, and 110 +/- 6 ppm for 2 d after a 2-d familiarization period. Once trained to use the intercompartment tunnels, the mice made extensive use of the EPC, with each group making more than 2000 intercompartment movements during 48 h. Video recording verified the results of the automatic tracking system, which detected and correctly determined mouse location for 79% of the moves. The use of photosensors proved to be ineffective in recognizing individual mice. Although the EPC would benefit from refinement and further development, it simplified analysis of locomotion behavioral data. Results of the preference study indicated that the mice exhibited no clear preference for, or aversion to, any of the experimental concentrations of ammonia and that the mice clearly preferred the upper 2 compartments of the chamber over the lower 2 compartments. Further investigation should be conducted to verify these preliminary results and explore other preferences of laboratory mice for environmental conditions and resources.

The use of a GIS-based inventory to provide a national assessment of standing waters at risk from eutrophication in Great Britain.

A three-tiered, hierarchical, risk-based prioritisation system was developed to assess the number of standing waters in Great Britain (GB) at risk from eutrophication. The scheme is based on four properties: importance, hazard, sensitivity to enrichment and sensitivity to recovery. Lake size, conservation status and legislative requirements were used to assess importance. The anthropogenic total phosphorus (P) load estimated from land cover, livestock and population data was used as a measure of the eutrophication hazard. Lakes with a retention time >3 days were considered to be sensitive to enrichment. The Wederburn depth (an estimate of the average summer thermocline depth) was used to predict the potential response of a lake to nutrient reduction. Lakes which were mainly stratified or fully stratified during the summer were expected to respond quickly to remediation. An initial Tier 1 risk assessment was made for all standing waters in GB (approximately 14,300 with surface area greater than 1 ha), using the four parameters derived from nationally available, GIS-based data sources held in the GB Lakes Inventory. Of the 2362 important lakes in GB, the system identified 1736 with low hazard but under potential threat because of their high sensitivity to enrichment. The system assessed that the ecology of 212 was likely to be damaged by eutrophication owing to high hazard and high sensitivity but with relatively poor chance of recovery following remediation. A further 332 lakes were considered to be damaged but were likely to respond to rehabilitation. In summary, the risk-based prioritisation system performed well and provides a useful tool for assessing standing waters at risk of eutrophication on a national basis. Inevitably, however, the need for nationally available datasets at Risk Tier 1 results in data resolution issues and errors may occur. The results highlight the importance of data validation using lake-specific information at Risk Tier 2.

Microbiological monitoring in individually ventilated cage systems.

Housing rodents in IVC racks has many advantages over conventional cages but also presents unique challenges related to health monitoring. The authors review the issues to consider in design of a sentinel program using IVC systems.

Efficacy of three microbiological monitoring methods in a ventilated cage rack.

The use of individually ventilated caging (IVC) to house mice presents new challenges for effective microbiological monitoring. Methods that exploit the characteristics of IVC have been developed, but to the authors' knowledge, their efficacy has not been systematically investigated. Air exhausted from the IVC rack can be monitored, using sentinels housed in cages that receive rack exhaust air as their supply air, or using filters placed on the exhaust air port. To aid laboratory animal personnel in making informed decisions about effective methods for microbiological monitoring of mice in IVC, the efficacy of air monitoring methods was compared with that of contact and soiled bedding sentinel monitoring. Mice were infected with mouse hepatitis virus (MHV), mouse parvovirus (MPV), murine rotavirus (agent of epizootic diarrhea of mice [EDIM]), Sendai virus (SV), or Helicobacter spp. All agents were detected using contact sentinels. Mouse hepatitis virus was effectively detected in air and soiled bedding sentinels, and SV was detected in air sentinels only. Mouse parvovirus and Helicobacter spp. were transmitted in soiled bedding, but the efficacy of transfer was dependent on the frequency and dilution of soiled bedding transferred. Results were similar when the IVC rack was operated under positive or negative air pressure. Filters were more effective at detecting MHV and SV than they were at detecting MPV. Exposure of sentinels or filters to exhaust air was effective at detecting several infectious agents, and use of these methods could increase the efficacy of microbiological monitoring programs, especially if used with soiled bedding sentinels. In contemporary mouse colonies, a multi-faceted approach to microbiological monitoring is recommended.

Shotgun identification of protein modifications from protein complexes and lens tissue.

Large-scale genomics has enabled proteomics by creating sequence infrastructures that can be used with mass spectrometry data to identify proteins. Although protein sequences can be deduced from nucleotide sequences, posttranslational modifications to proteins, in general, cannot. We describe a process for the analysis of posttranslational modifications that is simple, robust, general, and can be applied to complicated protein mixtures. A protein or protein mixture is digested by using three different enzymes: one that cleaves in a site-specific manner and two others that cleave nonspecifically. The mixture of peptides is separated by multidimensional liquid chromatography and analyzed by a tandem mass spectrometer. This approach has been applied to modification analyses of proteins in a simple protein mixture, Cdc2p protein complexes isolated through the use of an affinity tag, and lens tissue from a patient with congenital cataracts. Phosphorylation sites have been detected with known stoichiometry of as low as 10%. Eighteen sites of four different types of modification have been detected on three of the five proteins in a simple mixture, three of which were previously unreported. Three proteins from Cdc2p isolated complexes yielded eight sites containing three different types of modifications. In the lens tissue, 270 proteins were identified, and 11 different crystallins were found to contain a total of 73 sites of modification. Modifications identified in the crystallin proteins included Ser, Thr, and Tyr phosphorylation, Arg and Lys methylation, Lys acetylation, and Met, Tyr, and Trp oxidations. The method presented will be useful in discovering co- and posttranslational modifications of proteins.