**2. Journey through the COVID-19 pandemic phases in India and its impact on various sectors**

## **2.1 The initial impact**

Although the initial impact of the virus was seen majorly in the developed countries, India got affected during January when the first case of it was reported from the Kerala province in southern India [5].

Further, lockdown and curfew were imposed in different countries, which eventually dwindled the economy globally affecting the poor and middle-class earners, maximally. In India, the daily wagers were forced to flee from their working place to go back to their native places. Since transportation was at standstill due to lockdown these workers along with their family members had to cover their long distances on foot only. Sooner or later, many social activists took the initiative to help these sufferers. At this point, the blame game of the politicians towards each other started pouring in.

Over time, the COVID-19 pandemic has affected many sectors which majorly include health, education, hospitality, and entertainment sectors. Diagramatic representation as documented in **Figure 1**.

### **2.2 Impact on the health sector**

The situation on the other hand, in the hospital sector, was far more frightening as more and more casualties were happening due to limited beds and a greater number of patients requiring hospitalisation. Urgent steps were thought of and many

**Figure 1.** *Various sectors affected by the COVID-19 pandemic.*

### *A Journey through and beyond the COVID-19 Pandemic in Indian Setup-Lessons Learnt So Far DOI: http://dx.doi.org/10.5772/intechopen.100290*

places like hotels, train bogies, etc. were soon made as alternatives to hospital beds. Health care workers despite all the stress of contracting the disease themselves took the front seat to take care of the sufferers, but to their very surprise many people, by this time, had become prey to different myths and false beliefs and had to face the resistance from these society members. The situation was grave in certain parts of the country and police had to take some harsh steps to control the nuisance.

Amongst the patients, the effect was seen majorly in elderlies and in patients suffering from cancer and with comorbidities. In their article published by Wang et al., concluded that patients above 70 years of age had a faster progression of disease in the elderlies [6]. Similarly, a study was done by Liang et al. documented that case-fatality rates were higher in elderlies and also the pulmonary complication requiring resuscitation were higher in those suffering from cancer [7]. Besides its impact on these patients treatment delays due to the lockdown became a major issue. Various modifications in the treatment guidelines were framed by this time by various eminent treating societies to make the treatment continued and minimise the delays. Also, working protocols of different departments were modified and policies to screen the COVID-19 patients were formulated.

Though modified protocols were important from the clinical point of view, the psychological effect of the suffering had a deep impact on society. Effective healthrelated communication has thus remained a need of the hour. An article published by A. Finset et al. concluded that effective health communication is important in fighting the COVID-19 pandemic [8]. Similar importance of health communication has been documented by Reddy et al. in their article [9]. Amongst the various modes of communication, telecommunication with patients has also proved to a boom during the pandemic. As the majority of the patients who had queries related to the disease and treatment-related queries were also taken care of. Also, scheduling of all appointments further made things organised both for the patients as well as the treating physicians.

From a treatment point of view the rat race to treat and cure the ailment was soon catching the pace by this time. Many different permutation and combinations of drugs were thought of during this phase, with not much success to it. Alternative medicines like homoeopathy and ayurvedic soon joined the race and tried their best to come up with a possible solution. Since the disease was due to the virus, the only hope for it was thought of by generating an appropriate vaccine. Research on making the vaccine soon began. Different countries to the very best of their capabilities are trying till date, to make an effective vaccine. In India, Bharat Biotech took the initiative and completed the initial phases of the trials.

Covaxin and Covishield are the two vaccines that were developed and brought forward for mass vaccination. The Government decided to carry out the vaccination in a phased manner with age group 45 and above along with health care workers and patients with comorbidities receiving the initial jab. Although a majority of people have got vaccinated, a lot many still doubt the efficacy of the vaccine and remain concerned with its side effects. The false rumours regarding the vaccine have further added to it and have prevented a lot of many to refrain from getting vaccinated. While people were being told about the advantages and necessity of getting the vaccination done, the second wave of the pandemic came into existence.

### **2.3 Impact of the COVID-19 on the education sector**

The very initial phase of the COVID-19 pandemic deeply impacted the education front, as the lockdown and curfew forced all the schools, colleges, and universities to shut down. The digital model was soon implemented and the majority of the teaching soon began through online mode. An article published by Pravat Kumar Jena,

has highlighted the various initiatives taken by the Govt. Of India on the education sector. Also various positive and negative effects have been emphasised [10].

Although the effect of this form of education can only be assessed with time as of now, the digital mode has surely taken care of the education front.

### **2.4 Impact of the COVID-19 on the tourism and hospitality sector**

Besides the health and the education sector, the hospitality industry has also been affected badly by the pandemic. The hospitality industry accounts for 10% of the global GDP. The travel restrictions have not only led to revenue losses also the jobs of various workers have come to stake. As per United Nations World Tourism Organisation that international tourists will be down by 20–30%

In India, the hospitality industry is likely to be hit hard. Experts suggest that domestic hotel companies faced a weak FY20 and a much weaker FY21. The challenges and learnings from the COVID-19 pandemic in the Indian setting have been elaborated in the article published by V. Kaushal et al. [11].

Other than India, the major impact of the COVID-19 pandemic can also be seen in countries whose economy depends largely on the tourism and hospitality sector.

### **2.5 Impact of the COVID-19 on the entertainment sector**

Music and film plays a very important role in one's life and is not only a source of entertainment but is also a major revenue generator for the country. The lockdown and curfew have led to the downfall in film production and indirectly to revenue collection. G. Nhamo et al. in their study found that the pandemic has led to the cancellation of productions, films, and music festivals leading to multibillionaire losses which furthermore has led to a devastating impact on the livelihood of artists [12].

### **2.6 COVID-19 and international health security**

United Nations brought the concept of health security during the year 1994 [13]. Some of the alternative terms include global public health security, public health security, and global health security [14–18]. Global Health Security is defined as the activities required, both proactive and reactive, to minimise the danger and impact of acute public health events that endanger people's health across geographical regions and international boundaries [19]. Four well-known international organisations that takes care of the health security issues are- World Health Organisation (WHO), the United Nations (UN), the European Union (EU), and the Asia-Pacific Economic Cooperation (APEC).

A well-developed Health security system is the backbone of a strong infrastaructure of every country. The ongoing COVID-19 pandemic has shown how segregated and poorly funded health systems are worldwide. In a simplified form, it could be said that it has exposed the ground realities of the health security systems of all countries, revealing that no country is safe. Even countries like the USA, UK which receives top ratings for pandemic preparedness in the Global Health Security Index, have reported a significant number of COVID-19 cases and deaths [20]. India on the other hand, ranks at 57th position, with a cumulative score of 46.5, and fairs poor in the Global Health security index [21]. Compared to other countries, though the COVID-19 outbreak occurred late in India, the impact of it was much more severe. Further, the effect was seen in a wavy fashion with the first wave soon followed by the second wave. The impact of the second wave was much more severe than the first wave and majorly affected the younger population. Although India shares with

### *A Journey through and beyond the COVID-19 Pandemic in Indian Setup-Lessons Learnt So Far DOI: http://dx.doi.org/10.5772/intechopen.100290*

the United Nations the Global Health Security Agenda (GHSA), still a lot many steps are needed to be taken to improvise the Health security status.

Furthermore, not always security concerns are overlapped with health issues. Recently, both bioterrorism and emerging infectious diseases have raised alarming bells on health security issues [22–24]. An international cooperation is thus highly solicited in containing the spread of infectious diseases and enforcing a strong health security system. Also, establishing a global centralised Disaster Management Society and developing a comprehensive Global Disaster Rescue Plan could be possible measures for curtailing future pandemics.


*Article* **Coupling and Coordination Relationship between Urbanization Quality and Ecosystem Services in the Upper Yellow River: A Case Study of the Lanzhou–Xining Urban Agglomeration, China**

**Wusheng Zhao 1 , Peiji Shi 1,\*, Ya Wan 1 and Yan Yao 2**

**Abstract:** The study of the man–land relationship in the urbanization process is the current frontier and focus of international research. How to balance urban development and ecosystem conservation in the Upper Yellow River is a key issue for sustainable development in China. In this study, we evaluated the Lanzhou–Xining urban agglomeration (LXUA) by constructing a multi-dimensional assessment system for urbanization quality and ecosystem services. The efficacy function model, entropy weight method, and Integrated Valuation of Ecosystem Services and Tradeoffs (InVEST) model were used to quantitatively assess the subsystems' state of development. Then, the coupling model (CD) and the coordination degree (CCD) model were used to explore the coupling coordination relationship and spatial-temporal change characteristics of the composite system. The findings indicate that: (1) In 2020, the quality of urbanization in the LXUA showed the pattern of "double core". The development of urban centers in each city is insufficient, and the proportion of counties with a low level is too high. (2) Integrated ecosystem services showed an increasing distribution pattern from the northeast to the southwest. Water provision services, soil conservation services and carbon fixation services all showed growth trends. (3) Each county's composite system was in the run-in stage or highly coupled stage. The subsystems were closely related to each other. (4) The CCD was decreased by 6% between two decades. The number of counties on the verge of disorder was the highest. About 80% of the counties were relatively lagging behind in ecosystem services.

**Keywords:** urbanization quality; ecosystem services; coupling coordination; spatial-temporal variations; Lanzhou–Xining urban agglomeration


*<IP_ADDRESS> Preoperative examination and patient selection*

Some significant parameters that should be taken into account during *preoperative examination* for the patient selection for mIOL implantation and, then, for the selection of the appropriate type and power of the IOL are the following:

**107**

*Pseudophakic Presbyopic Corrections*

*DOI: http://dx.doi.org/10.5772/intechopen.96528*

corneal astigmatism into account [94].

with time through the process of neuroadaptation [99].

• *Visual axis, angle kappa:* In pseudophakic presbyopic corrections with implantation of mIOLs, the angle kappa (misalignment between the visual and pupillary axis) [100] should be taken into consideration, especially for hyperopic patients with a large angle kappa [101, 102]. It has been found that large angle kappa is correlated with more glare and halos after implantation of mIOLs [86, 103]. This happens because when decentration of mIOLs is present (intraoperatively or postoperatively), which is more common in eyes with large angle kappa, the light rays pass through a multifocal ring instead of the central optic zone, resulting in glare and halos [104]. Decentration higher than 0.75 mm

non-mIOL pseudophakic presbyopic correction when an ultrasound biometry is used for the IOL power calculation. The surgeon should discuss with the patient the risks caused by the measurement inaccuracy, and may choose an alternative

• *Corneal astigmatism:* Patients undergoing multifocal IOL implantation may not tolerate residual astigmatism of > 1 D or even 0.75 D according to some studies, and a toric multifocal IOL may be required in such cases [92]. For the best possible calculation of the corneal astigmatism, the performance of more than one keratometry technologies is suggested, since no perfect method is available [13]. Ideally, rotary prism technology (auto-keratorefractometers), PCI, corneal Scheimpflug tomography (which also takes into account posterior corneal astigmatism) or Placido-based corneal topography, and keratometry taken by image-guided systems for lens extraction surgery could be used [16]. In case of agreement in corneal cylinder power and axis among the utilized technologies, the surgeon can safely choose the appropriate common cylinder power of the toric IOL and the alignment axis. If the different methods of measuring corneal cylinder produce inconsistent results, the treatment of the ocular surface should be considered and the measurements should be repeated. In addition, the proper patient positioning should be confirmed [93]. Online calculators are also available for the toric/multifocal toric IOL power calculation with very good results because most of them take IOL power and posterior

• *Pupillary size:* Pupil size, but also pupil position in relation to the near and distance zones of the lens, seem to affect the optical performance of the implanted mIOLs in terms of VA, optical aberration, diffraction, and retinal luminance. Patients with wide pupils in scotopic light conditions who were implanted with mIOLs have a higher risk of poor postoperative contrast sensitivity under mesopic illuminance levels and optical phenomena such as glare, halos, and starbursts [95]. In fact, large pupil size is one of the most common causes of dissatisfaction and photic phenomena in patients with mIOLs [96–98]. Therefore, the pupillary size under photopic and mesopic light conditions before mydriasis must be determined preoperatively to minimize subjective postoperative side effects [95]. Everyday clinical practice has shown that a mesopic pupil size smaller than 5 mm can minimize photic phenomena. In case of pupil size larger than 5 mm, the surgeon should discuss with the patient the risks for dysphotopic phenomena and decide whether to proceed with the multifocal IOL implantation or not depending on the patient's visual needs and lifestyle. An alternative surgical solution could be proposed if multifocal IOLs are excluded. Nevertheless, in all cases regardless of the pupil size, patients should be informed about the possibility of appearance of some optical phenomena which, however, may be reduced

surgical solution according to patient's visual needs and lifestyle.

*Current Cataract Surgical Techniques*

from mIOL implantation [13, 61].

ruled out from insertion of mIOLs.

*<IP_ADDRESS> Preoperative examination and patient selection*

results in long eyes [82, 85].

vision [64]. In addition, patients who drive at night for long periοds of time

• *Occupation:* Patients with high occupational visual demands such as pilots and commercial (public service vehicle, taxi, or truck) drivers should be avoided

• *Expectations*: Hypercritical patients with unrealistic expectations are not suitable candidates for mIOL implant. For instance, people whose main concern is the sharpest clearest vision and who do not mind wearing near glasses even when reading very small letters or under low lighting conditions, should be

The surgeon should never promise full spectacle independence, but they should explain to patients that there is a good chance that they will not need glasses for the majority of their activities of daily living and that no perfect IOL to simulate their pre-presbyopic continuous vision exists yet [61]. However, spectacles may be needed under highly demanding conditions or during reading under low lighting conditions. Additionally, patients should be counselled about the optimal level and direction of light for easier reading. Finally, patients should be counselled about adverse events including halo, glare, reduced contrast sensitivity as well as discussing neuroadaptation in greater detail [75]. If patients are able to understand all benefits and risks of mIOLs, they could be good candidates to continue the preoperative examination for a potential pseudophakic presbyopic correction with mIOL

Some significant parameters that should be taken into account during *preoperative examination* for the patient selection for mIOL implantation and, then, for the

• *Optical biometry:* Careful biometry is crucial to accurate IOL power calculation and astigmatism management, since inaccurate biometry is the most common cause of residual refractive error postoperatively [76]. Third generation formulas such as the Holladay [77], SRK/T [78] and Hoffer Q [79] provide good outcomes for patients with average AL (22 – 25 mm) and keratometry. In cases of short eyes (AL < 22 mm) with a shallow ACD (< 2.40 mm), formulas that take preoperative ACD into account, such as Haigis, or alternatively Hoffer Q, could be used [80–82]. When dealing with long eyes (AL > 25–26 mm), SRK/T (with optimized constants), is still an accurate solution [83]. Haigis (with optimized constants) is accurate especially for eyes with AL < 30 mm [84]. Finally, new formulas such as Barrett Universal II and Olsen provide good

• *Dense cataract:* In case of dense cataracts when optical biometry measurements have low accuracy, the repetition of measurements could increase the accuracy in IOL power calculation. When no measurements can be taken, A-scan ultrasound biometry can be used. However, since the accuracy of this method is lower than the accuracy of optical biometry, [86–88] and taking into account that even a residual refractive error of 0.50 D, especially in mIOL implantation, can reduce the vision quality and increase photic phenomena, [89–91] the surgeon should be cautious with the choice of the mIOL or even with the decision for a possible

selection of the appropriate type and power of the IOL are the following:

should not be considered as good mIOL candidates.

**106**

implantation.

non-mIOL pseudophakic presbyopic correction when an ultrasound biometry is used for the IOL power calculation. The surgeon should discuss with the patient the risks caused by the measurement inaccuracy, and may choose an alternative surgical solution according to patient's visual needs and lifestyle.

irrespectively of IOL design has shown both far and near visual function deterioration [96, 105]. Clinical practice and research has shown that inclusion of patients with an angle kappa and angle alpha < 0.5 mm, ideally < 0.3 mm, (primarily in vertical and secondarily in horizontal axis in the Cartesian coordinates plot) both in mesopic and photopic conditions of illumination can minimize the risk for mIOL decentration and postoperative photic phenomena [103]. On the other hand, when angle kappa and angle alpha are > 0.5 mm, multifocal IOLs should be avoided, and monofocal IOLs for binocular distance vision or monovision could be chosen [106].

	- It is assumed that corneas which have undergone a refractive surgery such as laser in situ keratomileusis (LASIK), photorefractive keratectomy (PRK) or radial keratotomy (RK), have been rendered multifocal and show many

**109**

*Pseudophakic Presbyopic Corrections*

*DOI: http://dx.doi.org/10.5772/intechopen.96528*

powers are used [64, 118].

aberrations by the laser procedure. As a result, it also assumed that the implantation of a mIOL would further deteriorate the visual function.

○ An additional difficulty is the inaccuracy in IOL power calculation that comes from the inaccuracy in the determination of the total corneal refractive power, [64, 117] and the inaccuracy in the estimation of the effective lens position by various IOL power calculation formulas when post-laser corneal

○ Patients having corneas with irregular astigmatism, or more than 1 micron of higher corneal aberrations (HOA), especially if they are caused by high levels of coma, are not good candidates for mIOLs [64]. In addition, eyes with a large pupil (> 4 mm) have more possibilities to appear photic phenomena caused by existing aberrations in comparison with eyes with a small pupil (< 4 mm). As a result, a poorer visual quality under mesopic or glare conditions may occur [119].

○ Although mIOLs are usually well tolerated and effective after corneal refractive surgery, refractive surprises can be common, especially in patients with myopia greater than 6.0 D or in those that have undergone LASIK [120, 121]. To increase the refractive predictability and reduce the risk of a suboptimal refractive outcome, the precise IOL power calculation is of paramount importance. Since keratometers are unable to measure accurately K values of the central post-laser cornea and the outer and inner corneal surfaces may change unpredictably after corneal refractive surgeries, a variety of IOL power calculation methods have been suggested [64]. The most accurate method is the *clinical history method*. To apply this method and calculate the central corneal power, the refractive error (spherical equivalent) and the K values prior to the keratorefractive surgery as well as the stable refractive error after the surgery must be available [122, 123]. Apart from the numerous methods and formulas that are available, some online calculators can also be

used complementarily for the IOL power calculation [124–126].

implantation of the mIOL [130].

*3.2.3 Surgical technique*

○ In general, patients that had a previous keratorefractive surgery and want to have an additional pseudophakic presbyopic correction with mIOLs have high expectations for spectacle-free good vision [127]. However, they should always be informed for the possibility of inaccurate measurements, residual refractive error, hyperopic shift, and/or aberrations resulting in photic phenomena especially in mesopic light conditions [128]. Additionally, the surgeon should explain to patients that there is a possibility that they will need to use a miotic agent postoperatively, especially in mesopic and scotopic light conditions, in case of photic phenomena, [129] and that an additional corneal refractive surgery or even a further surgery for exchanging the mIOL may be needed in the future [130]. Ideally, if the corneal surface is irregular with corneal aberrations and a second laser treatment is necessary to correct this irregularity, such a treatment should be better performed prior to the

Phacoemulsification is the gold standard technique for cataract surgery and refractive lens exchange. Since mIOL implantation requires high precision and various factors should be taken into consideration, in addition to the common ones for the conventional cataract surgery with monofocal IOL implantation, a variety

*Current Cataract Surgical Techniques*

preferred [34].

vision or monovision could be chosen [106].

the visual and optical quality [111].

pre- and/or postoperatively [112, 115, 116].

irrespectively of IOL design has shown both far and near visual function deterioration [96, 105]. Clinical practice and research has shown that inclusion of patients with an angle kappa and angle alpha < 0.5 mm, ideally < 0.3 mm, (primarily in vertical and secondarily in horizontal axis in the Cartesian coordinates plot) both in mesopic and photopic conditions of illumination can minimize the risk for mIOL decentration and postoperative photic phenomena [103]. On the other hand, when angle kappa and angle alpha are > 0.5 mm, multifocal IOLs should be avoided, and monofocal IOLs for binocular distance

• *Pupil center shift (PCS):* It has been found that not only the photopic kappa angle, but also the PCS is associated with dysphotopsia after mIOL implantation [102]. In case of patients having a PCS higher than 0.4 mm, surgeons should decide whether the optical zone of mIOL should be centered on the photopic or mesopic/scotopic pupil center. In fact, when interpolated (from photopic 2 mm to scotopic 7 mm pupil) PCS is higher than 0.7 mm, the implantation of mIOLs should be avoided and other types of IOLs should be

• *Aberrations – ocular scattering:* Although aberrations, such as comma, SA or first order astigmatism, contribute to the enlargement of the depth of focus, they can also result in a decrease in contrast sensitivity and quality of vision. For instance, it has been found that anterior corneal coma values higher than 0.32 or 0.33 mm may create intolerable photic phenomena when a diffractive mIOL is implanted, thus contraindication of mIOL implantation has been suggested in higher coma values [13, 107, 108]. Additionally, since angle kappa [109] and PCS [110] can influence ocular aberrations, they should also be co-evaluated with ocular aberrations in order to provide patients with the best possible vision quality. Apart from aberrations, ocular scattering may have an impact on the quality of retinal image, which may be overestimated when only aberrations are taken into account. Therefore, measurement of both aberrations and ocular scattering could contribute to a more accurate assessment of

• *Dry eye disease:* Since dry eye disease and cataract are very common in the elderly population, but also the ocular surface and tear film play a significant role in the quality of vision, dry eye disease treatment should be considered as an integral part of the pre- and postoperative patient's management. It is well known that dry eye may reduce the vision quality after mIOL implantation [112]. Additionally, it has been observed that cataract surgery is also responsible for causing dry eye disease or aggravating existing dry eye symptoms [113, 114]. Therefore, artificial tears and eyelid hygiene, but also cyclosporine or autologous platelet-rich plasma (PRP) in more severe cases, should be used

• *Previous corneal refractive surgery:* Particular attention should be given to

ering a pseudophakic presbyopic correction with mIOL implantation.

patients who have undergone a prior corneal refractive surgery and are consid-

○ It is assumed that corneas which have undergone a refractive surgery such as laser in situ keratomileusis (LASIK), photorefractive keratectomy (PRK) or radial keratotomy (RK), have been rendered multifocal and show many

**108**

aberrations by the laser procedure. As a result, it also assumed that the implantation of a mIOL would further deteriorate the visual function.


**Meet the editor**

Marcelo Saad graduated in Medicine from the Federal University of Sao Paulo, Brazil, from where he also received his MSc and PhD in Rehabilitation Sciences. As medical specialties, he is board certified both in Physiatry (Physical Rehabilitation) and Acupuncture. Currently, he is President of the Medical Spiritist Association of S. Paulo and Director Member at the Medical College of

Acupuncture of S. Paulo. Besides his work as a physician, he develops activities in scientific publication, is on the editorial boards of some medical journals, delivers technical lectures, and participates in scholar tasks. His fields of interest are acupuncture, rehabilitation, complementary therapies, and spirituality in health.


**Abstract**

Forensic DNA analysis is widely used to determine kinship and the identity of evidence from the crime scene and it is especially important in the identification of human remains after different types of exposure (water, heat, etc.). Currently, there are no official recommendations for forensic scientists as to which bones and tissues are the most reliable among degraded DNA samples. Since 2014 more than 350 fragments of unidentified corpses have been examined in the Forensic Biological Department (Republic Bureau of Forensic Medicine, Kazan, Russia). Based on our experience, the most reliable and reproducible DNA profiles are obtained from lower limber bones (in 90% cases), muscles (in 85% cases) and ribs (in 80% cases). However, we discovered a new source of DNA – the odontoid process of the 2nd cervical vertebra, which contains a high amount of DNA with a better state of preservation than many other bones. According to our results, when a complete skeleton or unidentified corpse is found, it is advisable to provide bones with soft tissue remnants in the absence of deeply embedded putrefactive changes. When working at the crime scene, special attention should be paid to separating small bones and fragments from skeletal remains.

**Keywords:** Forensic genetics, degraded DNA, DNA profiling, autosomal and Y-chromosome STRs, mitochondrial DNA


**4. The roles executed by NK cells with molecules in cytotoxic granules, the cell surface ligand and secreted proteins**

As mentioned above, the cell surface expression of activating receptors on NK cells is a key event which defines the performance of those cells. What precisely occurs in NK cells following stimulation with KARs? NK cells execute two different events following recognition of target cells with activating receptors (**Figure 3**). The first event exerts natural cytotoxicity against the targets by releasing perforin and granzymes in cytotoxic granules into the space of the immune synapse between NK and target cells. Perforin is thought to function in generating a pore in the plasma membrane as complement proteins, and then granzymes enter through the pore and mediate apoptosis by deploying their serine protease activity [34]. Alternatively, NK cells also express cell surface FasL, which can also induce apoptosis through Fas receptors on the target cells [35]. Moreover, TNF-related apoptosis-inducing ligand (TRAIL) is also produced by NK cells and induces apoptosis of target cells like FasL [36–38]. This natural cytotoxicity itself highlights the importance of early removal of abnormal cells, which transiently appear in the body, and directly contributes to preventing the development of tumor diseases. However, it also has another role linked to antigen-specific cytotoxicity by cytotoxic T lymphocytes (CTLs) in acquired immunity. CTLs have to be primed by dendritic cells (DCs) with a complex comprising antigen peptide and MHC class I molecule before they can become effective cytotoxic cells from naïve cells. In this priming, extracellular antigen is exceptionally presented on MHC class I for CTLs by a particular subset of DCs, in a process referred to as "cross-presentation" [39–41]. These type of DCs endocytose dead target cells, digest their antigens and express a complex of MHC class I and antigen peptide on the cell surface [42], and the dead cells can be provided from early injury of target cells by NK cells to DCs [43]. That is why NK cells are linked to acquired

2B4 was low in those asbestos-exposed cells. Moreover, we examined the characteristics of human primary NK cells in PBMCs cultured with asbestos and found a decrease in cell surface NKp46 in patients with malignant mesothelioma, a tumor disease caused by inhalation of asbestos, and also showed impaired natural cytotoxicity. Less information is known about the natural ligands of NKp46. However, a previous investigation demonstrated that cell surface expression levels of NKp46 were correlated with the natural cytotoxicity of K562 and that reverse antibody-dependent cell-mediated cytotoxicity (ADCC) of P815 was correlated with antibodies to NKp46 [33]. Additionally, our previous study demonstrated that NK cells in healthy individuals with high natural cytotoxicity showed high expression of NKG2D, NKp46 and phosphorylation of ERK following stimulation *via* those receptors, whereas NK cells in individuals with low natural cytotoxicity showed the converse [2]. These results led us to surmise that determination of the gene expression level of activating receptors might be one important parameter in estimating the natural cytotoxicity of effector cells such as PBMCs *in lieu* of employing methods involving incu-

**4. The roles executed by NK cells with molecules in cytotoxic granules,**

As mentioned above, the cell surface expression of activating receptors on NK cells is a key event which defines the performance of those cells. What precisely occurs in NK cells following stimulation with KARs? NK cells execute two different events following recognition of target cells with activating receptors (**Figure 3**). The first event exerts natural cytotoxicity against the targets by releasing perforin and granzymes in cytotoxic granules into the space of the immune synapse between NK and target cells. Perforin is thought to function in generating a pore in the plasma membrane as complement proteins, and then granzymes enter through the pore and mediate apoptosis by deploying their serine protease activity [34]. Alternatively, NK cells also express cell surface FasL, which can also induce apoptosis through Fas receptors on the target cells [35]. Moreover, TNF-related apoptosis-inducing ligand (TRAIL) is also produced by NK cells and induces apoptosis of target cells like FasL [36–38]. This natural cytotoxicity itself highlights the importance of early removal of abnormal cells, which transiently appear in the body, and directly contributes to preventing the development of tumor diseases. However, it also has another role linked to antigen-specific cytotoxicity by cytotoxic T lymphocytes (CTLs) in acquired immunity. CTLs have to be primed by dendritic cells (DCs) with a complex comprising antigen peptide and MHC class I molecule before they can become effective cytotoxic cells from naïve cells. In this priming, extracellular antigen is exceptionally presented on MHC class I for CTLs by a particular subset of DCs, in a process referred to as "cross-presentation" [39–41]. These type of DCs endocytose dead target cells, digest their antigens and express a complex of MHC class I and antigen peptide on the cell surface [42], and the dead cells can be provided from early injury of target cells by NK cells to DCs [43]. That is why NK cells are linked to acquired

bation with NK-sensitive target cells.

186 Natural Killer Cells

**the cell surface ligand and secreted proteins**

**Figure 3.** Summarized illustration of the roles executed by NK cells following the recognition of targets with activating receptors and subsequent responses in acquired immunity. (1) NK cells recognize target cells with activating receptors including NKG2D and NKp46, which trigger the machinery for target cell injury. (2) NK cells exert the action of killing targets by perforin/granzymes, FasL or TRAIL. (3) Dead target cells are endocytosed as antigen (Ag) by DCs. (4) The particular subset of DCs which have endocytosed lysed target cells play a role in "cross-presentation" to stimulate naïve CD8<sup>+</sup> T lymphocytes to develop into mature CTLs. (5) The stimulated NK cells also produce cytokines including TNF-α and IFN-γ, (6)–(8) which stimulate DCs to produce IL-12 and other proinflammatory cytokines, thereby promoting Th1 cell polarization and CTL development.

immunity as well as function in innate immunity by themselves. Secondly, the production of cytokines including TNF-α and IFN-γ by NK cells after recognition of targets also plays an important role in DC maturation [43, 44]. Those cytokines stimulate DCs to produce IL-12 and other proinflammatory cytokines, which promote Th1 cell polarization and CTL development with specificity against target cells. Additionally, IFN-γ produced by NK cells is able to effect Th1 polarization directly. Those findings relating to the production of cytokines by NK cells, in particular, as IFN-γ is a key cytokine which is produced by NK cells and supports tumor immunity, leads us to hypothesize that the production of IFN-γ by NK cells in an individual might be utilized alone to estimate the performance of natural cytotoxicity in those cells. However, it is known that NK cells can be divided into two populations comprising CD56bright and CD56dim cells, which show different natural cytotoxicity and production of IFN-γ. CD56bright NK cells have high production of IFN-γ and low natural cytotoxicity, whereas CD56dim NK cells have low production of IFN-γ and high natural cytotoxicity [45, 46]. Those findings demonstrate that measurement of IFN-γ production by NK cells is insufficient to estimate the natural cytotoxicity of those cells and that determination of multiple parameters related to NK cells is necessary in order to effectively evaluate the performance of natural cytotoxicity in an indirect manner. All of this information indicates that the performance of NK cells is reflected by the strength of stimulation through cell surface activating receptors as well as the subsequent production of functional molecules as described above.