Immunobiology of COVID-19: Mechanistic and therapeutic insights from animal models

The distribution of the immune system throughout the body complicates in vitro assessments of coronavirus disease 2019(COVID-19)immunobiology,often resulting in a lack of reproducibility when extrapolated to the whole organism.Consequently,developing animal models is imperative for a comprehensive understanding of the pathology and immunology of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)infection.This review summarizes current progress related to COVID-19 animal models,including non-human primates(NHPs),mice,and hamsters,with a focus on their roles in exploring the mechanisms of immunopathology,immune protection,and long-term effects of SARS-CoV-2 infection,as well as their application in immunoprevention and immunotherapy of SARS-CoV-2 infection.Differences among these animal models and their specific applications are also highlighted,as no single model can fully encapsulate all aspects of COVID-19.To effectively address the challenges posed by COVID-19,it is essential to select appropriate animal models that can accurately replicate both fatal and non-fatal infections with varying courses and severities.Optimizing animal model libraries and associated research tools is key to resolving the global COVID-19 pandemic,serving as a robust resource for future emerging infectious diseases.

Evaluation of a rabbit rectal VX2 carcinoma model using computed tomography and magnetic resonance imaging

AIM： To establish a rabbit rectal VX2 carcinoma model for the study of rectal carcinoma.METHODS： A suspension of VX2 cells was injected into the rectum wall under the guidance of X-ray fluoroscopy. Computed tomography （CT） and magnetic resonance imaging （MRI） were used to observe tumorgrowth and metastasis at different phases. Pathological changes and spontaneous survival time of the rabbits were recorded.RESULTS： Two weeks after VX2 cell implantation, the tumor diameter ranged 4.1-5.8 mm and the success implantation rate was 81.8%. CT scanning showed low-density loci of the tumor in the rectum wail, while enhanced CT scanning demonstrated a symmetrical intensification in tumor loci. MRI scanning showed alow signal of the tumor on T1-weighted imaging anda high signal of the tumor on T2-weighted imaging.Both types of signals were intensified with enhanced MRI. Metastases to the liver and lung could beobserved 6 wk after VX2 cell implantation, and a largearea of necrosis appeared in the primary tumor. The spontaneous survival time of rabbits with cachexia and multiple organ failure was about 7 wk after VX2 cell implantation.CONCLUSION： The rabbit rectal VX2 carcinoma model we established has a high stability, and can be used in the study of rectal carcinoma.

Establishment of VX2 breast carcinoma model in rabbit by injecting tumor mass suspension

Objective: To establish a stable model of VX2 breast carcinoma in rabbit and select the optimal way. Methods: Thirty female New Zealand rabbits were randomly divided into 3 groups with 10 in each. Tumor cell suspensions or tumor mass suspensions were injected into breast tissues of rabbits of group A and B, respectively. Tumor blocks were surgically implanted in rabbit breasts of group C. Tumor formation rate, tumor growth rate, and tumor-bearing survival time was compared, and the histological feature of tumor was observed. Results: Models were established conveniently and successfully in rabbits received injection of tumor mass suspensions. Tumor proliferated rapidly with the biological feature of squamous cell carcinoma. Conclusion: VX2 breast carcinoma model in rabbit was established successfully. Intramammary injection of tumor mass suspension is the best method.

COX-2 in liver,from regeneration to hepatocarcinogenesis:What we have learned from animal models?

The use of animals lacking genes or expressing genes under the control of cell-specific promoters has signifi cantly increased our knowledge of the genetic and molecular basis of physiopathology,allowing testing of functional hypotheses and validation of biochemical and pharmacologic approaches in order to understand cell function.However,with unexpected frequency,gene knockout animals and,more commonly,animal models of transgenesis give experimental support to even opposite conclusions on gene function.Here we summarize what we learned on the role of cyclooxygenase 2(COX-2) in liver and revise the results obtained in 3 independent models of mice expressing a COX-2 transgene specifi cally in the hepatocyte.Upon challenge with pro-inflammatory stimuli,the animals behave very differently,some transgenic models having a protective effect but others enhancing the injury.In addition,one transgene exerts differential effects on normal liver physiology depending on the transgenic animal model used.

Comparison of two different laparotomy methods for modeling rabbit VX2 hepatocarcinoma

AIM:To compare two different laparotomy methods for modeling rabbit VX2 hepatocarcinoma.METHODS:Thirty New Zealand rabbits were randomly divided into two groups:A and B.Group A was assigned a traditional laparotomy method(embedding tumor fragments directly into the liver with tweezers).Group B was subjected to an improved laparotomy method(injection of tumor fragments into the liver through a 15 G syringe needle).The operation time, incision length, incision infection rate, and mortality rate were compared between the two groups after laparotomy.Magnetic resonance imaging(MRI) was performed to evaluate tumor formation rates and the characteristics of the tumors 2 wk after laparotomy.RESULTS:The mean operation times for the two groups(Group A vs Group B) were 23.2 ± 3.4 min vs 17.5 ± 2.9 min(P < 0.05); the incision length was 3.3 ± 0.5 cm vs 2.4 ± 0.6 cm(P < 0.05); and the mortality rate after 2 wk was 26.7% vs 0%(P < 0.05); all of these outcomes were significantly different between the two groups.The incision infection rates in the two groups were 6.7% vs 0%(P > 0.05), whichwere not significantly different.MRI performed after 2weeks showed that the tumor formation rates in the two groups were 90.9%vs 93.3%(P>0.05).These rates were not significantly different between the two groups.The celiac implantation rate and abdominal wall metastasis rate in the two groups were 36.4%vs 13.3%(P<0.05)and 27.2%vs 6.7%(P<0.05),respectively,which were significantly different between the two groups.CONCLUSION:The tumor formation rates were not significantly different between the two methods for modeling rabbit VX2 hepatocarcinoma.However,the improved method is recommended because it has certain advantages.

Promising xenograft animal model recapitulating the features of human pancreatic cancer

BACKGROUND Multiple sites of metastasis and desmoplastic reactions in the stroma are key features of human pancreatic cancer(PC).There are currently no simple and reliable animal models that can mimic these features for accurate disease modeling.AIM To create a new xenograft animal model that can faithfully recapitulate the features of human PC.METHODS Interleukin 2 receptor subunit gamma(IL2RG)gene knockout Syrian hamster was created and characterized.A panel of human PC cell lines were transplanted into IL2RG knockout Syrian hamsters and severe immune-deficient mice subcutaneously or orthotopically.Tumor growth,local invasion,remote organ metastasis,histopathology,and molecular alterations of tumor cells and stroma were compared over time.RESULTS The Syrian hamster with IL2RG gene knockout(named ZZU001)demonstrated an immune-deficient phenotype and function.ZZU001 hamsters faithfully recapitulated most features of human PC,in particular,they developed metastasis at multiple sites.PC tissues derived from ZZU001 hamsters displayed desmoplastic reactions in the stroma and epithelial to mesenchymal transition phenotypes,whereas PC tissues derived from immune-deficient mice did not present such features.CONCLUSION ZZU001 hamsters engrafted with human PC cells are a superior animal model compared to immune-deficient mice.ZZU001 hamsters can be a valuable animal model for better understanding the molecular mechanism of tumorigenesis and metastasis and the evaluation of new drugs targeting human PC.

Lower respiratory tract samples are reliable for severe acute respiratory syndrome coronavirus 2 nucleic acid diagnosis and animal model study

Severe acute respiratory syndrome coronavirus 2(SARS-Co V-2) and coronavirus disease 2019(COVID-19) continue to impact countries worldwide.At present, inadequate diagnosis and unreliable evaluation systems hinder the implementation and development of effective prevention and treatment strategies. Here, we conducted a horizontal and longitudinal study comparing the detection rates of SARS-Co V-2 nucleic acid in different types of samples collected from COVID-19 patients and SARS-Co V-2-infected monkeys. We also detected anti-SARS-Co V-2 antibodies in the above clinical and animal model samples to identify a reliable approach for the accurate diagnosis of SARS-Co V-2 infection. Results showed that, regardless of clinical symptoms, the highest detection levels of viral nucleic acid were found in sputum and tracheal brush samples, resulting in a high and stable diagnosis rate. Anti-SARS-Co V-2 immunoglobulin M(Ig M) and G(Ig G) antibodies were not detected in6.90% of COVID-19 patients. Furthermore,integration of nucleic acid detection results from the various sample types did not improve the diagnosis rate. Moreover, dynamic changes in SARS-Co V-2 viral load were more obvious in sputum and tracheal brushes than in nasal and throat swabs. Thus,SARS-Co V-2 nucleic acid detection in sputum and tracheal brushes was the least affected by infection route, disease progression, and individual differences. Therefore, SARS-Co V-2 nucleic acid detection using lower respiratory tract samples alone is reliable for COVID-19 diagnosis and study.

Experimental animal models of pulmonary hypertension:Development and challenges

Pulmonary hypertension(PH) is clinically divided into 5 major types, characterized by elevation in pulmonary arterial pressure(PAP) and pulmonary vascular resistance(PVR), finally leading to right heart failure and death. The pathogenesis of this arteriopathy remains unclear, leaving it impossible to target pulmonary vascular remodeling and reverse the deterioration of right ventricular(RV) function. Different animal models have been designed to reflect the complex mechanistic origins and pathology of PH, roughly divided into 4 categories according to the modeling methods: noninvasive models in vivo, invasive models in vivo, gene editing models, and multi-means joint modeling. Though each model shares some molecular and pathological changes with different classes of human PH, in most cases the molecular etiology of human PH is poorly known. The appropriate use of classic and novel PH animal models is essential for the hunt of molecular targets to reverse severe phenotypes.

Animal models for SARS-CoV-2 and SARS-CoV-1 pathogenesis,transmission and therapeutic evaluation

There is a critical need to develop animal models to alleviate vaccine and drug development difficulties against zoonotic viral infections.The coronavirus family,which includes severe acute respiratory syndrome coronavirus 1 and severe acute respiratory syndrome coronavirus 2,crossed the species barrier and infected humans,causing a global outbreak in the 21st century.Because humans do not have pre-existing immunity against these viral infections and with ethics governing clinical trials,animal models are therefore being used in clinical studies to facilitate drug discovery and testing efficacy of vaccines.The ideal animal models should reflect the viral replication,clinical signs,and pathological responses observed in humans.Different animal species should be tested to establish an appropriate animal model to study the disease pathology,transmission and evaluation of novel vaccine and drug candidates to treat coronavirus disease 2019.In this context,the present review summarizes the recent progress in developing animal models for these two pathogenic viruses and highlights the utility of these models in studying SARS-associated coronavirus diseases.

Establishing models of portal vein occlusion and evaluating value of multi-slice CT in hepatic VX2 tumor in rabbits

AIM： To establish models of portal vein occlusion of hepatic VX2 tumor in rabbits and to evaluate the value of multi-slice CT. METHODS： Forty New Zealand rabbits were divided into 4 groups according to digital table： Immediate group （group A; transplantation of tumor immediately after the portal vein occlusion）, 3-wk group （group B; transplantation of tumor at 3 wk after the portal vein occlusion）, negative control group （group C） and positive control group （group D）, 10 rabbits in each group. Hepatic VX2 tumor was transplanted with abdominalembedding innoculation immediately after the portal vein occlusion and at 3 wk after the portal vein occlusion. Meanwhile, they were divided into negative control group （Left external branch of portal vein was occluded by sham-operation, and left exite was embedded and inoculated pseudoly） and positive control group （Transplanted tumor did not suffer from the portal vein occlusion）. All rabbits were scanned with multi-slice CT. RESULTS： All 40 animals were employed in the final analysis without death. Tumor did not grow in both immediate group and 3-wk group. In 3-wk group, left endite was atrophied and growth of tumor was inhibited. The maximal diameter of tumor was significantly smaller than that in positive control group （2.55±0.46 vs 3.59±0.37 cm, t = 5.57, P 〈 0.001）. Incidences of metastasis in the liver and lung were lower in 3-wk group than those in positive control group （10% vs 400, and 90% vs 100%, respectively）. The expression intensities of the vascular endothelium growth factor （VEGF） in groups A, B, C and D were 0.10±0.06, 0.66±0.21, 0.28±0.09 and 1.48±0.32, respectively. VEGF expression level in the test group A was significantly lower than that in the negative control group C （t = 5.07; P 〈 0.001）.In addition, VEGF expression in the test group B was significantly lower than that in the positive control group D （t = 6.38; P 〈 0.001）. Scanning with multi-slice CT showed that displaying rate of hepatic artery branches was obviously lower in grade Ⅲ（40%） than that in grade Ⅰ（70%） and Ⅱ（100%） （P 〈 0.05）; but there was no significant difference in displaying rate of the portal vein at various grades. Values of blood flow （BF） of the liver, blood volume （BV）, mean transit time （MTT） and permeability of vascular surface （PS） were lower in the immediate group and 3-wk group than those in control groups, but values of hepatic arterial fraction （HAF） were increased. Significant positive correlations were existed between BF and BV （r = 0.905, P 〈 0.01）, and between BF and PS （r = 0.967, P 〈 0.01）, between BV and PS （r = 0.889, P 〈 0.01）. A significant negative correlation existed between PV and HAF （r = -0.768, P 〈 0.01）, between PS and HAF （r = -0.557, P 〈 0.01）. The values of BF, BV and PS had a positive correlation with VEGF （rBF = 0.842, rBV = 0.579, rPS = 0.811, P 〈 0.01） . However, there was no significant correlation between the values of MTT and HAF and the VEGF expression （rMTt = 0.066, rHAF = -0.027）. CONCLUSION： Ligating the left external branch of portal vein is an ideal way to establish models of portal vein occlusion in rabbits with hepatic VX2 tumor. Multi slice CT plays a key role in evaluating effect of portal vein occlusion.

Radiosensitivity of β-elemene on rabbit VX2 renal transplant carcinoma model

Objective To investigate the effects of low dosage of β-elemene on the radiosensitivity of rabbit VX2 renal transplant carcinoma model. Methods We took the rabbit VX2 renal transplant carcinoma as the model. Experimental rabbits were divided into three groups: the control group, the radiation group, and the radiation +β-elemene (radiosensitivity) group. The change of tumor was observed by Spiral CT and B ultrasound to compare its regrowth period. The tumor was measured by light microscopy and electron microscopy. Results The tumor in radiosensitivity group was restrained obviously and the sensitization enhancement ratio (SER) of β-elemene was 1.89. Different apoptosis was observed under transmission electron microscopy. Conclusion Low dosage β-elemene can enhance the radiosensitivity of rabbit VX2 renal transplant carcinoma model and induce the apoptosis of tumor cells, but the mechanism needs further study. It promotes apoptosis in mechanisms in vitro.

Characteristics of animal models for COVID-19

Severe acute respiratory syndrome coronavirus 2(SARS-CoV-2)is the causative agent of coronavirus disease 2019(COVID-19),the most consequential pandemic of this century,threatening human health and public safety.SARS-CoV-2 has been continuously evolving through mutation of its genome and variants of concern have emerged.The World Health Organization R&D Blueprint plan convened a range of expert groups to develop animal models for COVID-19,a core requirement for the prevention and control of SARS-CoV-2 pandemic.The animal model construction techniques developed during the SARS-CoV and MERS-CoV pandemics were rapidly deployed and applied in the establishment of COVID-19 animal models.To date,a large number of animal models for COVID-19,including mice,hamsters,minks and nonhuman primates,have been established.Infectious diseases produce unique mani-festations according to the characteristics of the pathogen and modes of infection.Here we classified animal model resources around the infection route of SARS-CoV-2,and summarized the characteristics of the animal models constructed via transnasal,localized,and simulated transmission routes of infection.

Retrieval,reporting and methodological characteristics for systematic reviews/meta-analyses of animal models:a meta-epidemiological study

The study aimed to analyze the reporting and methodological quality of systematic reviews(SRs)/meta-analyses(MAs)of animal models to provide references for later studies and avoid the waste of medical resources.EMBASE and MEDLINE databases were searched from inception to November 2017,with no language restriction.Two reviewers selected inclusion dependently and extracted the basic characteristics.Review Manager 5.3,stata 12.0,and SPSS 21 software were used to conduct analyses.A total of 46 SRs/MAs were included.The results showed that the English databases with high retrieval frequency are PubMed/MEDLINE,EMBASE,and Web of Science.67.31%(31/46)of the articles reported the search strategy in the full text or the appendix.65.22%(30/46)reported the literature screening flow diagram,and only 19.57%(9/46)reported the number of works of literature retrieved in each database.60.87%(28/46)illustrated supplement retrieval.Through 2 subgroup analyses,it was found that there were no significant differences in the quality of reports of PRISMA items.But referring to the methodological quality or reporting of PRESS items,SCI was better than that of non-SCI,while there seemed a source of funding to have no significant impact on the methodological quality or the items of PRESS.The results of PRESS,AMSTAR 2,and PRISMA were correlated,and the correlation between PRISMA and AMSTAR 2 was strong.These results demonstrated that search strategies of animal model SRs/MAs are still not enough comprehensive,report specification and methodological quality still need to be ameliorated.To show users the scientificity and rigor of the study,future research should focus on these various guidelines like PRESS,PRISMA,and AMSTAR 2 checklists that have been issued,it can help to increase the value of research and improve the utilization of medical resources.

Protective role of brain CYP2J in diverse Parkinson disease animal models

OBJECTIVE CYP2 family including CYP2C and CYP2J is the predominant arachidonic acid(AA)epoxygenase,and the epoxidation of AA produces four regioisomeric cis-epoxyeicosatrienoic acids(5,6-,8,9-,11,12-,and 14,15-EET).Human CYP2J2 is one of the main CYP isoforms expressed in brain,but CYP2C8 was present at a low level.The aim of this study is to investigate the roles of brain CYP2J in Parkinson disease.METHODS Rats received the right-unilateral y injection with concentrated LV-CYP2J3 or LV-EGFP in the substantia nigra(SN)at 3 d before LPS or 6-OHDA treatment.The animals were tested for rotational behavior with the dopaminergic agonist apomorphine dissolved in sterile saline at 14 and 21 d after LPS injection.The influence of CYP2J-dependent derivative,14,15-EET,on the genes related with oxidative stress was assayed in SH-SY5Y cells.RESULTS CYP2J overexpression or 14,15-EET treatment significantly increased the levels of SOD1,CAT,GPX1,NRF2 and KEAP1 in neurons.TLR4-My D88 signaling pathway was involved the down-regulation of CYP2J by LPS.The binding of p-CREB with the promoter of CYP2J was inhibited by the LPS treatment.The loss of dopaminergic neurons in the right SN induced by LPS or 6-OHDA was significantly decreased by CYP2J3 transfection at 21 d after LPS injection.Compared with LPS or 6-OHDA group,the number of the rotation of rats was decreased by 42.6% and 60.7%by CYP2J3 transfection at 14 d after LPS or 6-OHDA injection;meanwhile,the rotation number was decreased by 12.7%and 21.3%at 21 d.The accumulation of alpha synuclein induced by LPS was significantly decreased by CYP2J3 transfection.The mR NA levels of SOD1,CAT,GPX1,NRF2 and KEAP1 in SN were decreased by LPS,which was attenuated by the injection of LV-CYP2J3.CONCLUSION Brain CYP2J can play a protective role in the damage of the inflammation and oxidative stress to the dopaminergic neurons.Brain CYP2J-dependent derivatives from AA may have therapeutic effects in Parkinson disease via the up-regulation of the antioxidant system in neurons.

SARS-CoV-2 immunity in animal models

The COVID-19 pandemic,which was caused by severe acute respiratory syndrome coronavirus 2(SARS-CoV-2),has become a worldwide health crisis due to its transmissibility.SARS-CoV-2 infection results in severe respiratory illness and can lead to significant complications in affected individuals.These complications encompass symptoms such as coughing,respiratory distress,fever,infectious shock,acute respiratory distress syndrome(ARDS),and even multiple-organ failure.Animal models serve as crucial tools for investigating pathogenic mechanisms,immune responses,immune escape mechanisms,antiviral drug development,and vaccines against SARS-CoV-2.Currently,various animal models for SARS-CoV-2 infection,such as nonhuman primates(NHPs),ferrets,hamsters,and many different mouse models,have been developed.Each model possesses distinctive features and applications.In this review,we elucidate the immune response elicited by SARS-CoV-2 infection in patients and provide an overview of the characteristics of various animal models mainly used for SARS-CoV-2 infection,as well as the corresponding immune responses and applications of these models.A comparative analysis of transcriptomic alterations in the lungs from different animal models revealed that the K18-hACE2 and mouse-adapted virus mouse models exhibited the highest similarity with the deceased COVID-19 patients.Finally,we highlighted the current gaps in related research between animal model studies and clinical investigations,underscoring lingering scientific questions that demand further clarification.

Ferrets: A powerful model of SARS-CoV-2

The rapid spread of severe acute respiratory syndrome coronavirus 2(SARS-CoV-2) in recent years not only caused a global pandemic but resulted in enormous social,economic,and health burdens worldwide.Despite considerable efforts to combat coronavirus disease 2019(COVID-19),various SARS-CoV-2 variants have emerged,and their underlying mechanisms of pathogenicity remain largely unknown.Furthermore,effective therapeutic drugs are still under development.Thus,an ideal animal model is crucial for studying the pathogenesis of COVID-19 and for the preclinical evaluation of vaccines and antivirals against SARS-CoV-2 and variant infections.Currently,several animal models,including mice,hamsters,ferrets,and nonhuman primates(NHPs),have been established to study COVID-19.Among them,ferrets are naturally susceptible to SARS-CoV-2 infection and are considered suitable for COVID-19 study.Here,we summarize recent developments and application of SARS-CoV-2 ferret models in studies on pathogenesis,therapeutic agents,and vaccines,and provide a perspective on the role of these models in preventing COVID-19 spread.

A novel transgenic mouse model of Chinese CharcotMarie-Tooth disease type 2L

We previously found that the K141N mutation in heat shock protein B8 （HSPB8） was responsible for Charcot-Marie-Tooth disease type 2L in a large Chinese family. The objective of the present study was to generate a transgenic mouse model bearing the K141N mutation in the human HSPB8 gene, and to determine whether this K141NHSPB8 transgenic mouse model would manifest the clinical phenotype of Charcot-Marie-Tooth disease type 2L, and consequently be suitable for use in studies of disease pathogenesis. Transgenic mice overexpressing K141N HSPB8 were generated using K141N mutant HSPB8 cDNA cloned into a pCAGGS plasmid driven by a human cytomegalovirus expression system. PCR and western blot analysis confirmed integration of the KI41NHSPB8 gene and widespread expression in tissues of the transgenic mice. The K141N HSPB8 transgenic mice exhibited decreased muscle strength in the hind limbs and impaired motor coordination, but no obvious sensory disturbance at 6 months of age by behavioral assessment. Electrophysiological analysis showed that the compound motor action potential amplitude in the sciatic nerve was significantly decreased, but motor nerve conduction velocity remained normal at 6 months of age. Pathological analysis of the sciatic nerve showed reduced myelinated fiber density, notable axonal edema and vacuolar degeneration in K141N HSPB8 transgenic mice, suggesting axonal involvement in the peripheral nerve damage in these animals. These findings indicate that the KI4mHSPB8 transgenic mouse successfully models Charcot-Marie-Tooth disease type 2L and can be used to study the pathogenesis of the disease.

Single-nucleus transcriptomic profiling of multiple organs in a rhesus macaque model of SARS-CoV-2 infection

Infection with severe acute respiratory syndrome coronavirus 2(SARS-CoV-2) causes diverse clinical manifestations and tissue injuries in multiple organs.However, cellular and molecular understanding of SARS-CoV-2 infection-associated pathology and immune defense features in different organs remains incomplete. Here, we profiled approximately 77 000single-nucleus transcriptomes of the lung, liver,kidney, and cerebral cortex in rhesus macaques(Macaca mulatta) infected with SARS-CoV-2 and healthy controls. Integrated analysis of the multiorgan dataset suggested that the liver harbored the strongest global transcriptional alterations. We observed prominent impairment in lung epithelial cells, especially in AT2 and ciliated cells, and evident signs of fibrosis in fibroblasts. These lung injury characteristics are similar to those reported in patients with coronavirus disease 2019(COVID-19).Furthermore, we found suppressed MHC class I/II molecular activity in the lung, inflammatory response in the liver, and activation of the kynurenine pathway,which induced the development of an immunosuppressive microenvironment. Analysis of the kidney dataset highlighted tropism of tubule cells to SARS-CoV-2, and we found membranous nephropathy(an autoimmune disease) caused by podocyte dysregulation. In addition, we identified the pathological states of astrocytes and oligodendrocytes in the cerebral cortex, providing molecular insights into COVID-19-related neurological implications. Overall, our multi-organ single-nucleus transcriptomic survey of SARS-CoV-2-infected rhesus macaques broadens our understanding of disease features and antiviral immune defects caused by SARS-CoV-2 infection,which may facilitate the development of therapeutic interventions for COVID-19.

Expression Pattern of Testis-specific Expressed Gene 2 in Cryptorchidism Model and Its Role in Apoptosis of Spermatogenic Cells

In our previous study, we identified a novel testis-specific expressed gene 2 （TSEG-2） from mouse testis. To further investigate its functions, 35 male Balb/c mice （8 weeks old） were divided into cryptorchidism group （n=20）, sham group （n=10）, and control group （n=5）. In cryptorchidism group, the right testes were anchored to the inner lateral abdominal wall. In situ hybridization （ISH） was applied to measure the localization of TSEG-2 in mouse testis. Real-time quantitative PCR was performed to detect the expression of TSEG-2 gene. Meanwhile, under the mediation of polyethylenimine （PEI）, the recombinant vector pEGFP-TSEG-2 （n=5） or empty vector （mock, n=5） was transfected into the testis of male mice. The transfection efficiencies were measured under a fluorescence microscope. The apoptosis of spermatogenic cells was detected by terminal deoxynuleotidyl-mediated nick end labeling （TUNEL）. The results showed that TSEG-2 was expressed in convoluted seminiferous tubules, more precisely, in spermatogonia and spermatocytes. As compared with sham and control groups, the TSEG-2 transcription was significantly enhanced （P〈0.05） and was correlated with apoptosis of spermatogenic cells in cryptorchid testes （P〈0.05）. PEI was efficient in mediating transfeetion of TSEG-2 into seminiferous tubules of testis. One week post-transfection, intratesticular injection of TSEG-2 resulted in increased apoptosis of spermatogenic cells in vivo （P〈0.05）. These results indicate that TSEG-2 may participate in the apoptosis of spermatogenic cells and the pathogenesis of cryptorchidism.

5-Amino-4-oxopentanoic acid photodynamic diagnosis guided microsurgery and photodynamic therapy on VX2 brain tumour implanted in a rabbit model

Background Complete tumour resection is important for improving the prognosis of brain tumour patients. However, extensive resection remains controversial because the tumour margin is difficult to be distinguished from surrounding brain tissue. It has been established that 5-amino-4-0xopentanoic acid （5-aminolevulinic acid, ALA） can be used as a photodynamic diagnostic marker and a photosensitizer for photodynamic therapy in surgical treatment of brain tumours. We investigated the efficacy of ALA photodynamically guided microsurgery and photodynamic therapy on VX2 brain tumour implanted in a rabbit model. Methods Eighty New Zealand rabbits implanted with VX2 brain tumours were randomly assigned to five groups： control, conventional white light microsurgery, a photodynamic therapy group, a photodynamically guided microsurgery group and a group in which guided microsurgery was followed by photodynamic therapy. The VX2 tumour was resected under a surgical microscope. The tumour resection was confirmed with histological analysis. All animals were examined with MRI for presence of any residual tumour tissue. The survival time of each rabbit was recorded. Results All treatment groups showed a significantly extended survival time compared with the control group. Photodynamically guided microsurgery combined with photodynamic therapy significantly prolonged survival time, compared with guided microsurgery alone. MRI and the autopsy results confirmed removal of most of the tumours. Conclusions Our results suggest that photodynamically guided surgery and photodynamic therapy significantly reduce or delay local recurrence, increase the effectiveness of radical resection and prolong the survival time of tumour bearing rabbits. Their combination has the potential to be used as a rapid and highly effective treatment of metastatic brain tumours.