**3.2. Caspase independent pathway**

6 Apoptosis and Medicine

connect with other cell death pathway.

give us the direct impression about this type of apoptosis.

Fas-R/TNF-R Ligands

DISC

Caspase-8

Cytochorme C release

Apaf1

p53 induced protein

**Figure 1.** Summarize the caspase dependent apoptosis pathway

**ER**

Bax or Bid

Bcl-2 or Bcl-xL

p53

**Mitochondrion**

RFX

Radiation UV

Virus and bacteria infection

ROS

DNA demage

**Nucleus** 

from the bacteria and virus, we found that many researchers' data show that RNA fragments and DNA can also trigger caspase dependent apoptosis, such as RNA fragment produced by mycobacterium tuberculosis which in the early log-phase growth can trigger caspase-8 dependent apoptosis[3]; In vivo, DNA damage can trigger apoptosis through enhancing ROS level and changing the mitochondria membrane permeability; Many proteins or peptides will also make cell apoptosis, amyloid β peptide cytotoxicity can induce the intracellular calcium disturbance, and then the calpain will be activated by imbalanced calcium storage, While the calpain can activate caspase-12 which can located in ER to inactivate the Bcl-Xl, this is a novel caspase-12 dependent apoptosis pathway[4]. This way can be used by the mitochondria to

Above all, we can give the conclusion that pathogens, RNA or DNA, proteins or peptides, some chemical compounds or native compounds can all trigger caspase dependent apoptosis. They may have the different receptors and induce cell death through different caspase as the transducer to make the downstream signals transduction. The host used this way to defense the harmful factors and maintain the healthy physiological state. In the figure 1, we summarized the caspase dependent pathway transduction; we clearly known that the different ligands and receptors involved in this type of apoptosis, this picture will

FlASH knockdown

MCL-1 and short isoform of cFLIP

RIP1 TRIF

ER stress

release

containing death domain(PIDD)

Calcium

Calpain

Caspase-12

JNK activation Bax

ATP

DR4/5 CCL21/CCR7

dsRNA/polyI:C

TLR3

Caspase-9 Caspase-3 Substract cleavage

ROS

PIDDosome

Caspase-2

ERK Bcl-2

Caspase-3,Bax

apoptosis

Plasma membrane

Apoptosis have many mechanisms. It can be triggered by in vitro and in vivo cell ligands, and have different signal transduction pathways. Now, Apoptosis pathways are classified as caspase dependent pathway and caspase-independent pathway. In the above, we describe and conclude the caspase dependent pathway, which is characterized by the involvement of caspase in this type cell death process. In this paragraph, we will deep research the caspase independent apoptosis pathway. We will know that caspase does not participate in this type of process from the literally meaning. Up to now, this type of apoptosis has been founded by many researchers, and the research data give much information to us, this information can help us to understand the apoptosis complex mechanism, beside the mechanisms, the complicated ligands, which can induce this type of cell death, can also be found by many researchers. In the following, we will give some detailed contents about caspase independent apoptosis.

In the cell, a lot of ligands can induce mitochondria membrane potential change, the mitochondria damage will be the first step of the apoptosis, then ROS production increase, and ROS may the mainly factor to induce caspase independent apoptosis. For example, Denis Martinvalet found that granzyme A can directly induce the ROS increase and caspase independent mitochondria damage. Then the target of granzyme A, ER associated complex (SET complex), will translocated to nuclear and contributed to apoptosis [5]; AIF has been found the major important caspase-indenpendent pro-apoptosis factor, which can release from the mitochondria and translocate in the nuclear to cleavage the DNA, in the end, if the DNA damage have not been repaired by cells, the apoptosis will happen. Recently, many researchers found some compounds which can accompany with AIF production and induce cell death, such as simvastatin, staurosporine, cadmium and so on. These factors triggered caspase-independent PCD and fitted for the organism requirement; Beside AIF triggered caspase-independent PCD, ROS also participate in this type cell death. ROS can mediate poly (ADP-ribose) polymerase-1 (PARP-1) activation, and PARP-1 activation is necessary for AIF release from mitochondria. So ROS also involved in this type of cell death networks.[6].However, ROS participated in the caspase dependent apoptosis pathway also, Consequently, ROS might be the important bridge to connect two types of apoptosis in vivo. ROS mainly come from mitochondria, so mitochondria play important role in apoptosis pathways crosstalk. And the ligands usually trigger complicated reactions, including that AIF nuclear translocation, ROS increase and mitochondrial dysfunctions, these changes can cause to the caspase independent apoptosis pathway. For example, Cyclohexyl analogues of

Ethylenediamine Dipropanoic Acid, the compound that can induce peripheral blood mononuclear cells apoptosis of both healthy controls and leukemic patients through stimulating many apoptogenic factors activation(such as: AIF nuclear translocation, ROS increase and mitochondrial dysfunction ). [7] Referring the ROS, we propose that the GSH , NO, or other free radical groups may also take part in this type cell death, By this way, the GSH or NO modification proteins may also take part in the apoptosis pathway, GSH and NO can block some active thiol group and make the protein S-glutathionylation or Snitrosylation modification. These types modification may affect the protein's functions and make the cell to apoptosis result. Beside the AIF and ROS, there are many other ligands and signal molecular from the vitro or vivo cells as apoptogenic factors involving in caspase independent apoptosis pathway, such as lysosomal membrane permeabilization; some virus's protein; drugs; p53 suppression tumor factors or many other unknown compounds.

Extrinsic and Intrinsic Apoptosis Signal Pathway Review 9

In this type apoptosis, caspase family members did not involve in this cell death, and can not be inhibited by the caspase inhibitor {such as: the z-VAD-FMK; quinolyl valyl-o methylaspartyl[-2,6-difluorophenoxy]-methyl ketone(Q-VD-OPH);Ac-DEVD-FMK and so on}. In the cells, some components and events, such as the AIF; ROS; Ca2+; NAD+ and ATP;

Mitochondria, as a semiautonomous organelle in cells, apart from containing their own genetic material, play an important role in the energy metabolism. It can produce ATP to maintain the cell life activity and be known as an energy company in cells. Beside this major role, mitochondria are the places that the lot of biological reaction processes happened, such as ROS production, apoptosis, and regulation of aging [8] and so on. Mitochondria's dysfunction has the relation with many diseases (Alzheimer's disease; Parkinson's disease, cancer, diabetes) [9, 10]. These diseases have been identified to have some relation with the apoptosis; ROS produced by mitochondria have been regarded as one of important factors for apoptosis. Currently, many researches found the ROS produced increased when some pathogen infected, ROS can trigger apoptosis, through apoptosis, the pathogens may lost the perfect living environment, the host may defense the pathogen's diffusion by this manner. Due to these roles, mitochondria may be used as a proper therapeutic target to cure

As a dynamic organelle, mitochondria can change their shape and structure constantly to respond to the different stimuli and metabolic demands of cells. According to the latest researches about biochemistry and cell biology, the mitochondrial shape changes between fusion and fission play a very important role in the regulation of apoptosis [12, 13]. There were some debates about the opinion that apoptosis occurred relating with the mitochondria fission. These debates focused on which process happened firstly, either apoptosis is the result of the mitochondria fission and fragmentation, or reversely, as a following up affair, the mitochondria's fission and fragmentation happened at the downstream of apoptosis. While, we can sure that mitochondria shape dynamic changes

Calcium irons act as the upstream stimulus which can activate the cellular mitochondrial fission, the mitochondrial became fragmentation rapidly depended by the increased calcium level in intracellular. If the calcium level increased protractedly, mitochondria's fragmentation will be non-reversible and lead to apoptosis. So Jennifer R. Hom group regarded that the calcium involved in mitochondria morphology and participated in the apoptosis processing;[14] some mitochondria membrane proteins also have been found to control mitochondrial morphology, for example, the Bcl-2 protein resides in the outer mitochondrial membrane, and this protein acts as a central regulator of the intrinsic apoptotic cascade; while some other toxins or proteins can also regulate the mitochondrial fission/fusion, and these variation shapes of mitochondria was found to have relation with some diseases, For instance, the PD(Parkinson's disease) have been found with the

must be connected with apoptosis according to the follow latest researches:

protein misfolding and modification, can trigger the caspase independent apoptosis.

**3.3. Mitochondria dynamics and apoptosis** 

diseases related with this type cell death [11].

Up to now, the caspase independent apoptosis mechanism is still unknown clearly. Although some researchers have found AIF; ROS and other ligands can stimulate this type of PCD, the signal pathway still stay the phenomena stage and the deep mechanism should be dig out by us. No matter either apoptosis form, this type cell death has very important functions and deserves to be researched deeply. We collected a variety of information about this cell death and found that the caspase independent apoptosis existed in a mount of species and played indispensable role in cell growth, proliferation and death. In the figure 2, we give the outline about this type apoptosis pathway and the crosstalk manner between the different pathways, this picture will help us to know this type apoptosis well in the whole level.

**Figure 2.** Caspase independent apoptosis pathway

In this type apoptosis, caspase family members did not involve in this cell death, and can not be inhibited by the caspase inhibitor {such as: the z-VAD-FMK; quinolyl valyl-o methylaspartyl[-2,6-difluorophenoxy]-methyl ketone(Q-VD-OPH);Ac-DEVD-FMK and so on}. In the cells, some components and events, such as the AIF; ROS; Ca2+; NAD+ and ATP; protein misfolding and modification, can trigger the caspase independent apoptosis.
